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Much of today's biological research is motivated by the desire to understand higher-order processes more fully. For example, our desire to understand human development and disease helps us to drive research on cell biology. In turn, our desire to understand cells has motivated us to understand signaling pathways and gene networks. At each of these levels, networks — entities connected by arrows based on their relationships to one another — have proven to be a valuable way of representing knowledge (Figure 1).
Figure 1.
Example networks. (A) A signaling pathway with a cell (large green rectangle), proteins (colored ovals), a nucleus (dashed green circle), repression as blunted arrows, movement into the nucleus as a dashed arrow, and transcription as a solid black arrow. (B) The cell cycle with stages G1, S, G2, and M.Full figure and legend (18K)
Now our desire to understand the molecular underpinnings of biology and disease is motivating research into molecular kinetics. For example, it has recently been discovered that small oligomers of just a few Aβ peptides may be the toxic elements in Alzheimer's disease 1. Determining their structures could aid in designing drugs to prevent their formation, however, the structural heterogeneity of these oligomers makes accurate structural characterization with conventional methods difficult. Fortunately, computational modeling can capture both the dominant structures and dynamics of these molecules 2. Another example is proteomics. Now that genomics has given us the sequence of the human genome, there is a push for high throughput structure prediction to obtain the structures of all the proteins encoded therein. Information-based methods have proved useful for small globular proteins, but physical models (which capture both thermodynamics and kinetics) are likely required to push to systems like membrane proteins for which less structural data is available. Since the majority of drugs on the market target membrane proteins, such information would again be valuable for designing therapeutics.
As in many other fields, networks are a valuable framework for representing knowledge of molecular kinetics. In particular, networks called Markov state models (MSMs) are proving to be a powerful means of understanding processes like protein folding and conformational changes 3, 4, 5, 6, 7, 8, 9, 10, 11, 12. The power of MSMs derives from the fact that they are essentially maps of the conformational space accessible to a system. That is, like a road map with roads labeled with speed limits and cities labeled with populations, MSMs give the probability that a protein or other molecule will be in a certain set of conformations (called a state or node) and describe where it can go next and how quickly. Figure 2 shows a portion of an MSM as an example. Examining these maps can give tremendous insight into processes like protein folding and could even suggest how to manipulate these processes with small molecules, mutations or other perturbations. Models with sufficient resolution can also yield quantitative agreement with, or even prediction of, experimental observables like folding rates and structures 4, 5, 6.
Figure 2.
An example MSM for the villin headpiece. Shown here are four clusters of conformations automatically identified by MSMBuilder. Each cluster represents a state of the villin protein. Arrows indicate transitions between states, also identified by MSMBuilder. The group or cluster representing the native state (right-most) was accurately identified. Its members match the crystal structure (shown in darker blue and magenta) with an average root mean square deviation (RMSD) of 1.8 Å. Courtesy of Joy Ku and Gregory R. Bowman, reproduced with permission from Biomedical Computation Review.Full figure and legend (24K)
MSMs are typically constructed from simulation trajectories (i.e. series of conformations that were visited one after another in a physical simulation of a system, like a protein) 4, 5, 10, 12, 13, 14. Because of the temporal relationship between conformations in a trajectory, it is possible to group conformations that can interconvert rapidly into states and then determine the connectivity between states by counting the number of times a simulation went from one state to another. By employing these kinetic definitions, one ensures that the system's dynamics can be modeled reasonably well by assuming stochastic transitions between states 3, 4, 5, 6, 7, 10, 11, 13, 15. Thus, it is possible to perform analyses, such as identifying the most probable conformations at equilibrium. In addition, one can naturally vary the temporal and spatial resolution of an MSM by changing the definition of what it means to interconvert rapidly or slowly 4, 5, 15, 16, 17. By choosing a long timescale cutoff, one can obtain humanly comprehensible models with just a few metastable (or long-lived) states that capture large conformational changes, like folding. Such coarse-grained models are useful for gaining an intuition for a system. With a short timescale cutoff, on the other hand, one can obtain a model with many states. By using such high-resolution models, one sacrifices ease of comprehension for more quantitative agreement with experiments 4, 5, 18.
To date, MSMs have mostly been used to understand phenomena like peptide and protein folding 4, 5, 13, 17, 19, 20, 21, 22, 23, RNA folding 6, 16, and conformational changes 8, 9, 24. Having been validated by these studies, they are now being applied to important topics in human health. Examples include protein aggregation in Alzheimer's disease 2 and vesicle fusion 25, an important step in influenza infection.
The remainder of this review will be divided into four major sections. First, we review the application of MSMs to biomolecular folding, one of the driving problems behind the development of this technology. The next section focuses on the application of MSMs to human health, particularly protein misfolding diseases and influenza. This is followed by a review of MSM methodology and a discussion of efficiently capturing long timescales with short simulations and MSMs in the last two sections.
Protein and RNA folding
Protein folding is one of the biological problems that has driven the development of MSMs. From a biophysical point of view, it is simply amazing how proteins collapse to specific structures so quickly given the astronomical number of possible conformations they can adopt. From a biomedical point of view, understanding folding is an important first step towards understanding and preventing misfolding, as in Alzheimer's and Huntington's diseases 26. Better models for protein folding could also allow more efficient structure prediction 27, 28, aiding high-throughput proteomics initiatives.
Unfortunately, understanding protein folding is notoriously difficult. Much experimental work has been done to characterize protein folding, but no experiment can capture the dynamics of an entire system with atomic resolution in order to provide a complete picture of the process. Computer simulation has the potential to fill this void, but reaching biologically relevant timescales with sufficient (atomic) resolution is extremely challenging 29, and even with an infinitely long simulation one would still need a way to extract meaningful information from it. MSMs can overcome these issues by performing a kinetic clustering of conformations to provide maps of a system's conformational space and extracting long timescale dynamics from many short simulations 4, 5, 10, 13, 30, similar to the way relay runners can cover large distances despite the fact that each runner only goes a little way. As explained in the section 'Reaching biologically relevant timescales with MSMs', MSMs may even be used to direct simulations to where they are needed most (via adaptive sampling) to efficiently explore the conformations that a molecule can adopt 6, 31, 32, 33.
Many of the initial applications of MSMs have been to small peptides due to the challenges of simulating larger proteins 12, 13, 17, 20, 21, 22, 34, 35. For example, some of the fundamental methods were laid out by Singhal et al. 36 in an early work on a tryptophan zipper β hairpin. In another noteworthy example, Chodera et al.13 developed and applied one of the first automated methods for building MSMs to a number of small systems. Buchete and Hummer21 have also demonstrated how to tune MSMs to obtain models with varying degrees of coarse graining.
In one of the first forays into full protein systems, Jayachandran et al.37 constructed an MSM for the villin headpiece. Villin was selected because it has a hydrophobic core and tertiary contacts, which make it a real protein rather than just a peptide, but is still extremely small and fast folding 38, 39. Villin only has ~35 residues and folds on the microsecond timescale, whereas most proteins have a hundred or more residues and fold on millisecond to second timescales; thus, villin is more amenable to computer simulations, which are generally no longer than tens of microseconds.
Villin was also the first target to be studied by automated methods 4, mainly the MSMBuilder package 15, which is now freely available on the web (https://simtk.org/home/msmbuilder/). This new MSM, a portion of which is depicted in Figure 2, was validated by showing that it is capable of reproducing the raw simulation data. Moreover, it also yields reasonable agreement with experimental measurements. For example, based on free energies calculated from the model, it was possible to predict the native structure to within 2 Å of the X-ray crystal structure (Figure 2). This was a significant advance since the native state of a system is determined by the lowest free energy state, yet most structure prediction algorithms are still based on identifying low-energy conformations due to the difficulties inherent in calculating the entropic component of free energies 40, 41. Making predictions based on free energies will be especially important for more complex systems, like inherently disordered proteins, where the lowest energy state may not be the lowest free energy state. The longest timescales in our villin MSM were also consistent with experimentally measured folding rates 4. Capturing these rates is critical because most experimental observables can be derived given complete knowledge of the relevant rates and a few conformations from each state.
Similar methods have also been used to study the PinWW domain 5. This small protein consists of a three-stranded β-sheet and is another common model system for protein folding studies 42. One important contribution of this work was a method for probing the equilibrium ensemble of folding pathways, which is based on transition path theory 43 and related methods 44. This study also revealed numerous parallel folding pathways, as well as misregistered trapped states which slow the folding process.
MSMs have also been applied to the related problem of RNA folding 6, 16. This research has been driven by recent developments demonstrating that RNA does more than convey information from DNA to proteins and, like proteins, often must fold into specific three-dimensional shapes to accomplish these roles 45. The ribosome is a noteworthy example; however, it is far too large to sample exhaustively. Thus, simulations have tended to focus on small motifs, like RNA hairpins 46, 47. RNA hairpins are composed of a double helix stem capped by a short loop. Despite their small size, they play many important roles. They are ubiquitous in known RNA structures, may serve as nucleation sites for the folding of larger molecules and play important roles in RNA-RNA and RNA-protein interactions 46, 48. In the past, generalized ensemble simulations have been used to try to understand the thermodynamics of RNA hairpins 46, 49, 50. More recent work has begun to apply MSMs to capture both their thermodynamics and kinetics, showing that current force fields are at least sufficient to identify a hairpin as the native state and get the order of magnitude of the folding rate correct 6, 16.
Other studies have also used MSMs to try to understand phenomena like downhill folding 51, conformational changes in membrane proteins 9, the effects of mutations 52, proton pumping 53 and human disease (to be discussed in the next section). However, most of the work on protein and RNA folding has focused on methods development and validation. In the coming years, it will be exciting to identify common features in protein free energy landscapes, explain exactly how proteins are able to fold so quickly and elucidate the origins of experimental observables. Capturing longer timescales (i.e. milliseconds to seconds and beyond) will be crucial for these future developments. An important step in this direction is a recent work that was able to capture the folding of NTL9 54, an α/β protein that folds on the millisecond timescale 55. Further application to other biomolecules should also prove fruitful.
MSMs and human health
Protein folding is not the only process that is difficult to characterize experimentally. Inherent disorder in processes like aggregation and vesicle fusion also makes them good candidates for study with MSMs. Again, experimental studies of these systems have limited resolution. Computer simulations can provide atomic-level descriptions of their dynamics and, with MSMs, reach the relevant timescales.
One particularly noteworthy example of applying MSMs to human disease is a recent work on Aβ aggregation, which plays an important role in Alzheimer's disease 1, 2. Aβ is extremely difficult to work with experimentally; however, with computer simulations and MSMs, Kelley et al. 2 were able to obtain models with atomic resolution capable of capturing dynamics on tens of seconds timescales. To accomplish this feat, they first ran atomistic simulations of encounter complexes with varying numbers of monomers. An MSM was then constructed from this data using state definitions based on physical intuition (e.g. based on the number of monomers, dimers, trimers and tetramers). To achieve experimentally relevant concentrations of Aβ, they then added a diffuse state and rates of transitioning between this state and encounter complexes were calculated using an analytic diffusion theory. This model, a schematic of which is shown in Figure 3, was found to give reasonable agreement with experimentally measured rates, a tremendous achievement given that standard simulations are about six orders of magnitude shorter than the relevant timescales. The MSM also gave important mechanistic insight into Aβ aggregation. In particular, the authors identified a reasonably populated C-terminal β-hairpin that was the main source of interactions between aggregated peptides, leaving the N-termini exposed to the solvent. The exposure of the N-termini to solvent makes them more accessible for binding with other molecules, explaining why antibodies targeting the N-terminus tend to have higher binding affinities than those targeting the C-terminus.
Figure 3.
Markovian model for Aβ oligomerization. Our model was built using the different aggregation states as the Markov states; in a system with four chains, there are five such states: four monomers (MMMM), two monomers and one dimer (MMD), two dimers (DD), one monomer and one trimer (MT), and finally, one tetramer (Q). In addition, to include the effects of low concentration found experimentally, we discriminate EC states (in which states are close) from separated states. The rate-limiting steps in the aggregation process are shown as dotted lines. The numbers associated with the transitions are transition probabilities. The significant figures were determined from the uncertainties in the transition probabilities. Some transitions with very low probability have not been shown for the sake of clarity. Reprinted with permission from Kelley NW, Vishal V, Krafft GA, & Pande VS, J Chem Phys, 129, 214707, 2009. Copyright 2009, American Institute of Physics.Full figure and legend (50K)
In another example, MSMs were used to study vesicle fusion 25. Vesicle fusion is important for processes like infection by enveloped viruses (e.g. in influenza); however, their large size and slow timescales make them inaccessible to standard computer simulations. Typical simulations have ~10 000 atoms and reach tens of nanoseconds timescales, but understanding vesicle fusion requires reaching hundreds of microseconds timescales for systems with over 500 000 atoms. Once again, a combination of simulations with MSMs was able to overcome these limitations, leading to important mechanistic insights. First, vesicles were found to fuse via two pathways: a fast pathway wherein two vesicles become connected by a short stalk and then rapidly fuse, and a slow pathway in which the stalk state transitions to a strongly metastable hemi-fused state before finally fusing. In a later work, the authors were even able to probe the dependence of the relative probabilities and rates of these pathways on the lipid composition of the vesicles involved 56.
Other human diseases are also excellent candidates for study with MSMs. For example, thousands of short simulations have already been performed for the Huntingtin protein 57 and the influenza hemagglutinin fusion peptide 58. Thus, these systems are prime for study with adaptive methods 6, 31, 32, which allow one to build an initial MSM with available simulation data and then refine it by running new simulations from each state.
MSM methodology
The dynamics of proteins and other molecules are governed by the system's underlying free energy landscape. Much like hikers on a natural landscape, proteins prefer to stay in the valleys of their landscape and only rarely (and generally slowly) cross over the barriers and peaks between valleys. MSMs are essentially maps of such landscapes 3, 4, 5, 6, 7, 10, 13, 15, 19. However, whereas road maps have cities connected by roads labeled with speed-limits, MSMs have conformational states (sets of conformations in the same valley) connected by edges labeled by the probabilities of transitioning between them.
While MSMs may be visualized as networks, as in Figure 2, computers represent them as transition probability matrices (P) 3, 7, where the entry in row i and column j (Pij) gives the probability of transitioning from state i to state j in an interval called the lag time of the model (). The probability of being in any state at a particular time can be represented as a vector, v(t). The time evolution of v(t) can then be calculated using
where P() is a transition matrix with lag time and each multiplication advances the model through time by one lag time.
The eigenvalue/eigenvector spectrum of a transition probability matrix gives important thermodynamic and kinetic information 3, 7. For example, the first eigenvalue always has a value of one and the corresponding eigenvector gives the equilibrium probabilities of all the states. Subsequent eigenvector/eigenvalue pairs give information about sets of states that interconvert on the same timescale and the rates at which these transitions occur. For example, a rate can be calculated from an eigenvalue using
where μ is an eigenvalue, is the lag time, and k is a rate. This equation comes from the equivalence between discrete time MSMs and continuous time master equations (see Refs 10 and 13 for details). Using these properties and a few representative conformations from each state, it is possible to compute experimental observables 4, 5. One can even put error bars on these properties using Bayesian statistics 59, 60.
MSMs are straightforward to work with once you have a valid state definition (one yielding Markovian behavior). One simply has to assign conformations to these states and represent trajectories as a series of state assignments rather than as a series of conformations. One can then simply count the number of transitions between pairs of states and store these values as a transition count matrix (C), where the entry in row i and column j (Cij) gives the number of observed transitions from state i to state j. One can then obtain a transition probability matrix (P) by normalizing each row of the transition count matrix. From P, one can then calculate all the eigenvalues, eigenvectors and observables they desire.
The most challenging part of building MSMs is identifying a good state decomposition. A good state decomposition should group conformations that can interconvert rapidly into the same state because this implies that they are not separated by a significant free energy barrier. Conformations that cannot interconvert rapidly, however, should be separated into different states because they are likely separated by a significant free energy barrier.
Practically speaking, it is neither possible nor desirable to determine the rate (or equivalently the probability) of transitioning between two conformations. Rather, one must consider the rates of transitioning between sets of conformations. To understand this, one can imagine trying to measure how long it takes to get home from work. You could measure the time it takes to get from the front doorway of your office to the front doorway of your home. However, it would likely be more appropriate to measure the time it took you to get from anywhere in your work place to any point in your home, regardless of whether you left work through the stairwell door and entered your home through the front door or left work through the main door and entered your home through the garage door. In the same spirit, measuring the transition rates between sets of kinetically equivalent conformations is more meaningful than measuring the transition rates between individual conformations.
In the case of MSMs, one often obtains initial sets of conformations by clustering them into microstates based on geometric criteria (Figure 4), with the objective of having conformations within a given microstate be so similar that their geometric similarity implies a kinetic similarity 10, 13, 15. This initial decomposition may be ideal for making quantitative comparisons with experiments 4, 5, 18. To gain an intuition for the system, kinetically related microstates can then be lumped into macrostates to ensure a direct connection to the underlying free energy landscape (e.g. identify various valleys and how quickly one can get from one to another) 10, 13, 15. For example, in an earlier automated algorithm, one first generated a set of microstates, lumped them into macrostates using a method called PCCA 61, and then iteratively broke the macrostates up into new microstates and re-lumped them until convergence 13. A more recent automated algorithm (MSMBuilder), which is now freely available at https://simtk.org/home/msmbuilder/, avoids any iteration by using a different clustering algorithm to obtain finer clusters and then doing a single round of lumping using either PCCA or an improved method, called PCCA+ 62, 63. This procedure is outlined schematically in Figure 4.
Figure 4.
Schematic representation of the steps required for building an MSM and obtaining representative conformations for each state. First, Generalized Ensemble (GE) data (or other data for that matter) represented by points are grouped into microstates represented by circles, with darker circles for more highly populated microstates. Kinetically related microstates are then lumped together into macrostates or metastable states, represented by amorphous shapes. Finally, representative conformations are obtained by extracting the most probable conformation from each macrostate. Reprinted from Methods, 49, Bowman, GR, Huang, X, Pande, VS, Using generalized ensemble simulations and Markov state models to identify conformational states, 197-201, Copyright (2009), with permission from Elsevier.Full figure and legend (26K)
Regardless of how an MSM is built, it is necessary to choose a lag time and validate the final model. Checking that the implied timescales of the model level off as the lag time is increased is a first indication that a given state decomposition is reasonable and allows one to choose a lag time (the lag time at which the implied timescales first level off should be chosen) 64. Intuitively, this equates to checking that the slowest rates (often on the order of hundreds of nanoseconds or greater) are invariant with respect to the interval at which you count transitions (often on the order of ten nanoseconds or less). Unfortunately, evaluating where the implied timescales of a model level off is extremely subjective and new methods are needed to replace this criterion. Some new methods employing information theory and Bayesian statistics point to how this may be done 59, 65. Once a lag time has been selected, the transition probability matrix can be calculated and used to further validate the MSM by comparing its dynamics to the raw simulation data (the Chapman-Kolmogorov test) 4, 5, 13.
Another important question is how many macrostates to construct when building a model to gain some intuition for a system. In the past, researchers have chosen a number of macrostates based on gaps in the implied timescales 10, 13, 15. Doing so imposes a separation of timescales – fast intrastate transitions, slow interstate transitions – that should give rise to Markovian behavior, i.e., the next state a trajectory visits should be independent of its history because it should be able to enter a state and quickly lose memory of where it came from before transitioning to a new state. However, recent work has shown that there is often a continuum of timescales without any obvious gap. Thus, the number of macrostates has come to be seen as a tunable parameter that can be adjusted based on one's objectives 4, 5, 18. For example, to gain an intuition for a system one may desire to lump a dataset into as few states as possible, while still preserving the Markov property and reasonable agreement with the raw simulation data.
Reaching biologically relevant timescales with MSMs
An important advantage of MSMs is their ability to capture long timescale dynamics from many short simulations 4, 5, 9, 15, 21, 30. Traditional approaches to molecular simulation require global equilibration, i.e., each simulation must be orders of magnitude longer than the slowest relaxation process so that every possible transition can be observed multiple times 66. In contrast, MSMs only require local equilibration 6, i.e., each simulation only needs to be long enough to equilibrate within a subset of all the possible states. One can then obtain a global model by statistically stitching together many short, parallel simulations covering different parts of the conformational space, much like creating a quilt from many small patches or relay runners covering long distances.
Adaptive sampling algorithms take this reductionist approach a step further 6, 31, 32. In adaptive sampling, one first obtains an initial model and then uses Bayesian statistics to calculate the contributions of each state to statistical uncertainty in some observable of interest, like the rate of the slowest process. A new round of simulations is then started from these states in order to increase the model precision as efficiently as possible. Thus, one avoids running more simulations where they are unnecessary while gathering more data where it can be of most use. Work with toy models has shown that performing multiple rounds of adaptive sampling can quickly refine a model 31. Indeed, it has recently been shown that adaptive sampling can reduce the wall-clock time necessary to achieve a given model quality by a factor of N, where N is the number of parallel simulations run during each iteration 33. Adaptive sampling can also reduce the total computer time necessary for a given model quality by a factor of two 33. An exciting future direction will be to apply adaptive sampling to real systems of biological significance that are currently beyond the reach of computer simulations, like conformational changes in transcription elongation 67.
Even with adaptive sampling, however, MSMs are not without their limitations. For example, it is still unclear how to distinguish systematic errors from statistical uncertainty and correct for them too. Furthermore, while MSMs are excellent for describing free energy landscapes in terms of their basins, capturing transition states can also be quite important for understanding the mechanisms of conformational changes 46. Recent work on the application of topological methods to understanding free energy landscapes shows how to capture such transition states 46, 68. Indeed, progress on combining MSMs and topological methods to capture both free energy basins and transition states is already yielding insight into processes like RNA folding 16. Moderate speedups may also be achieved by building MSMs from generalized ensemble simulations 6, 11, 15, 69, which perform a random walk in temperature to take advantage of broad sampling at high temperatures to improve mixing between states at lower temperatures 70, 71.
Concluding Remarks
MSMs are a powerful means of mapping out the conformational space of both macromolecules and macromolecular complexes. While much of the literature on MSMs has focused on methods development and validation, they have already provided important insights into processes like protein folding, aggregation and vesicle fusion. Further progress in this direction will likely have important medical benefits, both by providing a deeper understanding of the molecular phenomena underlying higher-order biological processes (and especially human disease) and by allowing more effective drug and protein design. Methodological advances in the use of MSMs could also prove useful in systems biology and eventually find application in other fields relying on network representations, such as the study of gene networks and signaling pathways.
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- Chu VB, Herschlag D. Unwinding RNA's secrets: advances in the biology, physics, and modeling of complex RNAs. Curr Opin Struct Biol 2008; 18:305–314. | Article | PubMed | ChemPort |
- Bowman GR, Huang X, Yao Y, et al. Structural insight into RNA hairpin folding intermediates. J Am Chem Soc 2008; 130:9676–9678. | Article | PubMed | ChemPort |
- Koplin J, Mu Y, Richter C, Schwalbe H, Stock G. Structure and dynamics of an RNA tetraloop: a joint molecular dynamics and NMR study. Structure 2005; 13:1255–1267. | Article | PubMed | ChemPort |
- Uhlenbeck OC. Tetraloops and RNA folding. Nature 1990; 346:613–614. | Article | PubMed | ISI | ChemPort |
- Villa A, Widjajakusuma E, Stock G. Molecular dynamics simulation of the structure, dynamics, and thermostability of the RNA hairpins uCACGg and cUUCGg. J Phys Chem B 2008; 112:134–142. | Article | PubMed | ChemPort |
- Garcia AE, Paschek D. Simulation of the pressure and temperature folding/unfolding equilibrium of a small RNA hairpin. J Am Chem Soc 2008; 130:815–817. | Article | PubMed | ChemPort |
- Voelz VA, Luttmann E, Bowman GR, Pande VS. Probing the nanosecond dynamics of a designed three-stranded Beta-sheet with a massively parallel molecular dynamics simulation. Int J Mol Sci 2009; 10:1013–1030. | Article | PubMed | ChemPort |
- Muff S, Caflisch A. Kinetic analysis of molecular dynamics simulations reveals changes in the denatured state and switch of folding pathways upon single-point mutation of a beta-sheet miniprotein. Proteins 2008; 70:1185–1195. | Article | PubMed | ChemPort |
- Kim YC, Wikstrom M, Hummer G. Kinetic gating of the proton pump in cytochrome c oxidase. Proc Natl Acad Sci USA 2009; 106:13707–13712. | Article | PubMed
- Voelz VA, Bowman GR, Beauchamp KA, Pande VS. Molecular simulation of ab initio protein folding for a millisecond folder NTL9(1-39). J Am Chem Soc 2010; 132:1526–1528. | Article | PubMed | ChemPort |
- Horng JC, Moroz V, Raleigh DP. Rapid cooperative two-state folding of a miniature alpha-beta protein and design of a thermostable variant. J Mol Biol 2003; 326:1261–1270. | Article | PubMed | ChemPort |
- Kasson PM, Pande VS. Control of membrane fusion mechanism by lipid composition: predictions from ensemble molecular dynamics. PLoS Comput Biol 2007; 3:e220. | Article | PubMed | ChemPort |
- Kelley NW, Huang X, Tam S, et al. The predicted structure of the headpiece of the Huntingtin protein and its implications on Huntingtin aggregation. J Mol Biol 2009; 388:919–927. | Article | PubMed | ChemPort |
- Kasson PM, Pande VS. Predicting structure and dynamics of loosely-ordered protein complexes: influenza hemagglutinin fusion peptide. Pac Symp Biocomput 2007; 12:40–50.
- Bacallado S, Chodera JD, Pande V. Bayesian comparison of Markov models of molecular dynamics with detailed balance constraint. J Chem Phys 2009; 131:045106. | Article | PubMed | ChemPort |
- Noe F. Probability distributions of molecular observables computer from Markov models. J Chem Phys 2008; 128:244103. | Article | PubMed | ChemPort |
- Deuflhard P, Huisinga W, Fischer A, Schütte C. Identification of almost invariant aggregates in reversible nearly uncoupled Markov chains. Lin Alg Appl 2000; 315:39–59. | Article
- Deuflhard P, Weber M. Robust Perron cluster analysis in conformation dynamics. Lin Alg Appl 2005; 398:161–184. | Article
- Weber M, Kube S. Robust Perron Cluster Analysis for various applications in computational life science. Comput Life Sci Proc 2005; 3695:57–66.
- Swope WC, Pitera JW, Suits F. Describing protein folding kinetics by molecular dynamics simulations. 1. Theory. J Phys Chem B 2004; 108:6571–6581. | Article | ChemPort |
- Park S, Pande VS. Validation of Markov state models using Shannon's entropy. J Chem Phys 2006; 124:054118. | Article | PubMed | ChemPort |
- Rao F, Caflisch A. Replica exchange molecular dynamics simulations of reversible folding. J Chem Phys 2003; 119:4035–4042. | Article | ChemPort |
- Wang D, Bushnell DA, Huang X, et al. Structural basis of transcription: backtracked RNA polymerase II at 3.4 angstrom resolution. Science 2009; 324:1203–1206. | Article | PubMed | ChemPort |
- Yao Y, Sun J, Huang X, et al. Topological methods for exploring low-density states in biomolecular folding pathways. J Chem Phys 2009; 130:144115. | Article | PubMed | ChemPort |
- Muff S, Caflisch A. ETNA: equilibrium transitions network and Arrhenius equation for extracting folding kinetics from REMD simulations. J Phys Chem B 2009; 113:3218–3226. | Article | PubMed | ChemPort |
- Mitsutake A, Sugita Y, Okamoto Y. Generalized-ensemble algorithms for molecular simulations of biopolymers. Biopolymers 2001; 60:96–123. | Article | PubMed | ChemPort |
- Huang X, Bowman GR, Pande VS. Convergence of folding free energy landscapes via application of enhanced sampling methods in a distributed computing environment. J Chem Phys 2008; 128:205106. | Article | PubMed | ChemPort |
We thank AN Bowman (Department of Developmental Biology, Stanford Universtiy) for help with Figure 1. This work was funded by NIH R01-GM062868, NIH U54 GM072970, and NSF EF-0623664. GRB was supported by the NSF GRFP.
These links to content published by NPG are automatically generated.
REVIEWSThe role of dynamic conformational ensembles in biomolecular recognition
Nature Chemical Biology Perspective
RESEARCHSurrogate Endpoints for the Treatment of Venous Leg Ulcers
Journal of Investigative Dermatology Original Article


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Interventions and relations at Greenbrier are guided by the model's philosophy of relationality. The following concepts represent core elements of the therapeutic process. I probably incorporate all of these statements into my conversations on at least a weekly basis. Review these statements and see how many you in your life and with your interactions with others.
- All interventions at Greenbrier are reflective of mission, Source, and virtue.
- Mental health can be defined as the presence of virtue in all relationships.
- Virtue is the answer that will empower your destiny.
- Mission can simply be defined as taking the skills and attributes that you have been given and using them in a way that helps others.
- Source is the greatest thing greater than self.
- Relationality is much more than the connection between two individuals and includes:
- Intra-personal, i.e. the relationship we have with conflicting parts of self, should I or shouldn't I?
- Inter-personal, which includes friends and family, but also relationships that have yet to materialize, i.e. the relationship with a child that hasn't been born, with the man or woman that will become your husband or wife, etc.
- Relationship with Source.
- Relationship with nature.
- When we have helped a student, we have encouraged them to examine change on as many relational dimensions as possible.
How has Greenbrier Academy helped you to be more aware of mission, Source, and virtue in your life?
Mike Beswick, LICSW, BCD
Greenbrier Academy Counselor

subset: commoncrawl


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First of all, I want to apologize for the extra long writing hiatus I took. You know, life just gets in the way sometimes. If you're wondering what the hell I was up or where I was the whole time - I'm gonna try to distract you with this:
Got this goody bag from my sister on one of her NYC trips. Hmmm.. what's in it?
Clockwise from top left: Victoria's Secret "The Lacie" Thongs in Whipped Vanilla and Strawberry Delight, Skin Food Eye & Lip Remover, Libra Tampons (in very inconspicuous packaging), and Co. Bigelow Mentha Lip Tint in Magenta Mint.
Did you miss me?
Saturday, May 1, 2010
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Socialist Party councillor Mick Barry this morning defended the aims and actions of anti-property tax campaigners who forced the cancellation of last night's meeting of Cork City Council. He said: "The property tax is an unjust tax which will be used to fund the bank bailout and payments to bondholders. The campaigners sent a clear message to Fine Gael and Labour last night that this tax will be resisted and I will back that resistance all the way.".
Socialist Party Cork City Councillor defends anti property tax protest
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The Gaffer Pack for the Mole Richardson 575 watt HMI Par Kit includes basic accessories, such as:
CSR575 HMI Bulb Gloves with Mitt Clip AC Cube Tap, C-47's Assorted Gel Pack Ditty Bag


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I've been studying herbs since the 1970s…yes, just dated myself…and I've found several sites that give great information and detailed formulas. I purchase the herbal formula in bulk from FirMeadowLLC and have been using it for about 4 years now. I just this year finally made up some "cookies" for the dogs and goats but before last month I only dropped the herbs on their food and let them lick it up. Never had a dog or goat refuse to lick the bowl clean! When I made the 'cookies' for the goats they adored them (who wouldn't love something with molasses) but the two large livestock guardians (made with peanut butter base) refused to eat it so I had to close their mouths and make them swallow, but the two house dogs ate them like they were candy! So, I'll continue to sprinkle doses on top of the dogs food and since it's easier than making cookies will probably still top dress the goats feed as well, but I'll give cookies to the two little house dogs.
FirMeadowLLC – I order a lot of her formulas for my family and my animals.
To Sing With Goats – I recently found this site with a detailed recipe for making your own herbal wormer.
Fiasco Farms – I have this site bookmarked, I started making cheese with her instructions, and love the info she has on all aspects of goat raising.
Herbal Legacy – I've been a student of Dr. Christopher's since the 1970s, everything he has written is important to read for the knowledge it contains.


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Entire home/apt
5 guests
2 bedrooms
3 beds
About this listing
House 5238 in the town of Stomorska, Šolta - Central Dalmatia contains accommodation units of type Apartment (2) and is 320 m away from the sea. A beach is nearby (concrete slabs). The house is categorized as "Pensions". As the house is divided into several accommodation units, other guests will most likely be present during your stay. The hosts will not be in the house for the duration of your holiday. The house owner is under no obligation to accept additional persons and pets that were not stated in the reservation request and it is necessary to report them in advance.
The space
Apartment can accommodate 5 guests. Beds are situated in 2 sleeping rooms, but also in the living room, within 44m2. Guests can enjoy a sea view from the private terrace. Food can be prepared using barbecue which is shared with other guests too. This accommodation unit welcomes pets at an additional charge.
INCLUDED IN THE PRICE: tourist tax, final cleaning, parking, hairdryer.
OBLIGATORY ADDITIONAL COSTS: short stay surcharge.
OPTIONAL ADDITIONAL COSTS: breakfast (€4.00, a 50% discount for children aged 4 to 12 years, Free for children under 4 years), half-board (€10.00, a 50% discount for children aged 4 to 12 years, Free for children under 4 years), full board (€16.00, a 50% discount for children aged 4 to 12 years, Free for children under 4 years), laundry (€10.00 washing), boat mooring (€60.00 per night), pets (€5.00 per night), bicycle (€20.00 daily (per piece)).
The availability of additional services for your chosen dates of stay, as well as their prices (if none are listed) must be checked ahead of time. Any additional services not shown in the price calculation are payable directly to the property owner and only if used by the visitor.
Courtyard (50m2, Sitting area), Parking lot (Number of parking spaces: 3, Distance from the house: In the courtyard), Barbecue (shared), Breakfast, Half board, Full board, Laundry service, Boat mooring (Distance from the house: 320 m, Type of boat berth: shore), Bicycle (Quantity: 5), Hairdryer
Number of stairs from the property to the beach: 25
Car access possible: Yes
The facility is situated in relatively quiet surroundings
The property isn't surrounded by greenery
Sea: 320 m
Beach: 320 m
Supermarket: 560 m
Restaurant: 560 m
Emergency room: 10 km
Post office: 650 m
Disco: 10 km
Diving Centre: 550 m
Clinic: 10 km
Airport: 55 km
Pharmacy: 10 km
Exchange office: 650 m
Tourist info: 800 m
No of people can fit in is 5
Smoking. Pets allowed.
Super Strict 60
Cancel up to 60 days before your trip and get a 50% refund. Cancel within 60 days of your trip and the reservation is non-refundable.
No reviews (yet)
This home doesn't have any reviews. If you stay here, your review will appear here.
My name is Leana and I work for travel agency Adriatic . hr from Split, Croatia.
For the last 17 years we have been helping numerous travelers from all around the world to find their perfect accommodation on Croatian coast.
I know traveling to an unknown destination can be stressful, but with my help you will find yourself on a quiet island beach in no time. Maybe you prefer long summer beach parties instead? Let me know, and we will work together to find you exactly what you want.
Feel free to contact me!
Don't forget to check out our other Airbnb profiles for more destinations!


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Kingsport 20 QBG SE Series
The Kingsport Special Edition unit by Gulf Stream Coach offers a cozy space to enjoy your time with family and friends away from home.
Inside model 20QBG you will find a Grand View u-shaped dinette in the rear with a storage shelf on the road side. At night the dinette table can be folded down to provide additional sleeping space. There is an entertainment center nearby for your enjoyment.
The kitchen in this unit features a three burner range, double sink, refrigerator, and storage pantry for all of your canned and dry goods, plus more.
There is a convenient bathroom that is centrally located that includes a tub/shower, toilet, sink, and linen cabinet.
The front offers a queen size bed with wardrobes on either side, plus overheads storage, and so much more!Have a question about this floorplan?Contact Us
Sleeps 4 Length 24 ft 11 in Ext Width 8 ft Ext Height 11 ft 3 in Int Height 6 ft 8 in Interior Color Coastal, Seahawk, Night Sky, Mineral Hitch Weight 640 lbs Dry Weight 4600 lbs Cargo Weight 3040 lbs Fresh Water Capacity 51 gals Grey Water Capacity 37 gals Black Water Capacity 37 gals Furnace BTU 25000 btu
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Fun Town RV is not responsible for pricing errors, typos, or incorrect information on our website. Manufacturer and/or stock photographs, floor plans, and specifications may be used. Prices listed include dealer preparation, walk through orientation, and factory freight. Prices exclude sales tax, license fee, documentary fee, and state inspection fee. Any calculated payment information is an estimate only and does not constitute a guarantee that financing or a specific rate or term is available. Units are subject to prior sale until a buyers order is submitted and a deposit made. Please verify unit availability by calling 844-242-5100 as our inventory changes rapidly.


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READY IN: 20mins
Recipe by FLUFFSTER
I know all of you "baby boomers" will appreciate this, and you younger boomers will hopefully remember this recipe one day!
Top Review by Glory to God
Super quick and easy....kids loved it...it was delicious!
See Discussion:5 Reviews
Ingredients Nutrition
- 12 Hostess Twinkies
- 3 -4 bananas
- 1 (16 ounce) can crushed pineapple, undrained
- 1 (5 ounce) package instant vanilla pudding
- 12 ounces non-dairy whipped topping
Directions See How It's Made
- Slice Twinkies lengthwise, and line the bottom of a 13x9" pan.
- Slice bananas over Twinkies, followed by pineapple with juice.
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- Refrigerate.


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If you've just located the Church of Christ Women Authors Blog you may not know that this blog started July 15, 2015. I had already started the page and group Church of Christ Women Authors on Facebook, when someone suggested we have a catalog of books.
Well, I had no funds to make a catalog, but I knew I wanted to keep on helping all church of Christ women authors succeed, so I clicked on Blogger, created the blog, and I guess the rest is history. In the last couple of months this blog has become my baby, and I am so excited to watch it grow.
So, with that said, if you are a women author who is a member of the church of Christ, let me know and I will do a blog post about you and a link to your books. Also, keep an eye open for new authors and new books, as well as writing tips, articles and book sales.
Thanks again, and I am so happy to be able to talk to you every day through this blog!


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Gucci Pochette Authentic Pre Owned
This item is no longer available
Authenticity code: Gucci 00138122123This Gucci Pochette is classy and chic. It features black monogram canvas and a nice hand held strap. It is the perfect bag. Be the center of attention with this Gucci pochette!
Authentic: Pre Owned
Exterior: Gently used
Interior: Gently used
Measurements: 10L x 6H x 3W. Handle drop: 9 (Approx.)
Product Code: 104887
All photos are of actual item being sold. This Gucci® item is guaranteed to be authentic. Gucci is a registered trademark of Gucci. LuxeDH is not affiliated with or represents Gucci in any way or form.
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Here's What I Know About Web Design Inspiration 2016
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Here's What I Know About Web Design Inspiration 2016
The templates for Joomla are utilised to provide you with the total look of the webpage. Thus, it is better to steer clear of templates. By employing online ready-made templates, you don't have to be graphically trained to receive it done. You can begin with a completely free template that source online at no cost.
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Website plays an important function in this world, without a website nobody can get world great small business lead. As a result, in case you have designed your site couple of years back, you'll most probably require a latest technology inside your site. As a way to jazz up your website understand what you require and don't require.


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Navigation: Home | Alumni News | Alumni Events | Available Theses
Welcome to this Alumni page which we hope will be of particular interest to anyone who has completed the Diploma/ Postgraduate Diploma/ Postgraduate Certificate/MSt in Applied Criminology and Police Management or Police Studies.
We would like to use this page to help you maintain your links with us as well as update you on what is going or what may be relevant.
We will post any items you are happy to provide for public circulation job changes/publications/Alumni Events.
We hope as many of you as possible will come to the next International Conference on Evidence Based Policing. For further details see: Here
Annual Alumni Dinner
The 2017 Alumni dinner will be held at Fitzwilliam College on Weds 12th July for details please see: Here.
The 2018 Alumni Dinner will be held at King's College on Weds 11th July. Details will be posted in due course
Select from the categories below to filter the news
IoC Lee Centre wins VC Research Impact Award
We are happy to report that in an Oscar-Award setting in the University Combination room this Thursday, the announcer opened the envelope to say which of six nominees won the 2017 Vice-Chancellor's Research Impact Award for the School of Humanities and Social Sciences-- so that the Vice Chancellor could present the award and the 1,000 pound prize. And the winner was the Jerry Lee Centre for Experimental Criminology, for its development of the Cambridge Crime Harm Index! Hearty congratulations are due to the award recipients: Prof. Larry Sherman, Peter Neyroud, Eleanor Neyroud, Dr. Cristobal Weinborn and Dr. Barak Ariel.
More information cann be found here. (Scroll down for pictures of our award winners with the Vice-Chancellor.)
Review of the 10th Evidence-Based Policing Conference
A very positive of our 10th Annual Conference has been published here.
Geoffrey BarnesAlumni Update
Western Australia's new criminology head is focussing on innovative solutions to complex policing problems. Internationally renowned US crime expert, Geoffrey Barnes, has joined WA's police force as their inaugural Crime Director.
Listen to the interview here.
Alf Hitchcock
We are all very saddened to hear of the recent death of Alf Hitchcock, Chief Constable of the MOD following a short illness.
Our condolences go to all of his family and friends.
American Society of Evidence-Based Policing Conference
May 22-23 2017
The American Society of Evidence-Based Policing is hosting our first conference in partnership with the Police Foundation and Arizona State University. The conference is May 22-23, 2017 at Arizona State University, Phoenix. The conference gives practitioners a platform to present the research they have conducted or evidence-based practices they implemented in their organization. We have several areas of research being presented. The conference will be a robust 2 days of learning about research in policing.
As our first conference we are offering an extremely reasonable price - $150/ non-members and $75.00 members. I have attached a flyer and a tentative agenda. You can register for the conference or become a member at www.AmericanSEBP.com.Futher Information can be found here: Flyer, Agenda.
Australian National UniversityAlumni Update
We are delighted to report that Professor Sherman & Professor Heather Strang have been appointed to the post of Honorary Professor of the Australian National University School of Regulation and Global Governance (RegNet), College of Asia & Pacific. See Here
Debbie Platz
We are delighted to announce that Acting Assistant Commissioner Debbie Platz, MSt (Cantab) has accepted a position as Assistant Commissioner, National Manager Crime Operations with the Australian Federal Police.
Debbie was in Cambridge 2014-15, graduating in 2016 and has a long and distinguished career in the Queensland Police Service and I know you will join me in congratulating and wishing her well with her future career.
Interesting News
Shocking example of unintended consequences: Mandatory domestic violence arrests raise death rate 400%, study finds. Read the whole article at National Post
Ten One Article
"London Liaison Officer Superintendent Gary Smith attended Cambridge University's Evidence-Based Policing Conference to catch up on the latest developments – and also caught up with three officers studying at the university." Page 19
Download the excerpt here Ten One August 2014
Annual Update
Police Theses have been marked and presented to the Board of Examiners. Congratulations to the following who have been awarded distinctions: Nicole Cornelius (Hampshire Police), John Denley (West Midlands Police), Mike Hulett (SOCA), Brandon Langley (West Midlands Police) &Jo Rowland (Hampshire Police).
The Board are also pleased to announce that Mike Hulett will be awarded the Sir Richard Mayne Award for outstanding academic achievement on the M.St. in Applied Criminology and Police Management and has been invited to make a short presentation on his research at the 7th International Conference on Evidence Based Policing, 9 July 2014.
Congratulations to all who did so well.
There has been much in the news recently about UK police forces experimenting with the use of Body Worn Cameras. The original study was completed by Chief Tony Farrar from Rialto as part of his thesis for the MSt in Applied Criminology and Police Management
Please also see:
Police Foundation
Dana M. Gillis, Federal Bureau of InvestigationAlumni Update
After 27 years of federal government and military service, I retired in 2011 and have since taken a position
as Director of Athletic Compliance at Alabama A&M University, USA.
I work with collegiate athletes and coaching staffs to ensure compliance with National Collegiate Athletic Association policies and regulations. I'm around sport all-day, everyday. Really enjoying the opportunity to mentor young women and men as they prepare for life in the "real world". I also acquired a Level 5 Certificate in Executive Coaching and Mentoring from the Institute of Leadership and Management through the Footdown School of Coaching, Bath, UK.
Howard Veigas, Derbyshire PoliceAlumni Update
"Since graduation, my thesis was presented the EBP Symposium at GMU ( Washington) with Dr Lum in 2011.
Since then, I have been working on a co-authored article about the translating EBP research into practice, with Dr Lum. In 2013 we published our article. The reference is -
Veigas, H., and Lum, C. (2013).Assessing the evidence-base of a police service patrol portfolio.Policing.7(3), 248-262.
Frankie Taylor, PSNIAlumni Update
"I have been collaborating with Barak on publishing a systematic review into whether electronic monitoring has an effect on re offending rates. This has been accepted by the Campbell Collaboration Crime and Justice Group.
I am currently studying with London Metropolitan three years into a four year Doctorate programme with the John Grieve Foundation."
Karen Daber, Cambridgeshire PoliceAlumni Update
"I've leading on Police Integrity and just launched for public and service wide consultation the first ever code of ethics for policing in England and Wales."
Gilbert Houalla, Thames Valley PoliceAlumni Update
I am doing some work on my police area which may be of slight interest. I will try to explain it briefly:
Legislation in the UK requires the relevant Councils (County or Unitary) under the Safeguarding of Children agenda to carry out a 'return interview' with every child that goes missing. He vast majority of them will be high risk. The interview must be carried out by an independent organisation/person and the purpose of this return interview is to ascertain the reasons why that child went missing and if they were subject to any criminality such as sexual exploitation, drug offences and such like. This information will be used in any subsequent risk assessment. The evidence suggests that few authorities are carrying out return interview with every child, however return interviews are being carried out on children who consistently and persistently go missing.
I have managed secure finances to employ two staff members from Barnardo's to carry out return interviews on every child that goes missing in the County of Buckinghamshire but not Oxfordshire or Berkshire (three counties in TVP). The funding is for one year, the evaluation criteria has been agreed based on reducing children going missing and the potential of harm and as a consequence reduce the demand on policing."
Owen Weatherill, Hertfordshire PoliceAlumni Update
"I can report something positive already from my research. It has prompted some real interest in what information and data AVL (automated vehicle location) can provide and as a direct consequence we are now gearing up for a trial of extending this to officers personal radios, with a specific focus upon Neighbourhood policing. We are looking to use it to compare how the footprint of officers and PCSO's patrol compares for foot patrol vs those patrolling on cycles, when compared against demand data.
The backdrop to this is that our Chief invested in a fleet of cycles when he arrives around 2 years ago, but we just don't know what we get from this in real terms. Larry will be pleased to hear that I suggested that we use APL (Automated Personal location) to provide an evidence based assessment of the cycle usage, by monitoring officers' personal radio locations.
Its early days and won't start until mid November. It does however provide a very helpful element to my findings and conclusions section for my thesis, as this results directly from the work we have already done around analysing AVL data."
Page Header image Copyright to Martin Pettitt


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Are you ever bogged down by headaches, neck and shoulder pain due to stress and tension? Do you have muscle aches and pains in your back? I sure do.
I've been having a lot going on. I have been experiencing more depression and heartache and feeling the stress of my families every day needs. I know we all go through these things from time to time. It seems your body will take on the effects of things going on in your mind and heart.
With all the things going on here at my house I've been experiencing some major tension headaches. You know the ones coming from your shoulder blades, shoulders and/or neck going all the way up to the crown of your head.
When you have those headaches, body aches and pains what do you do to relieve the tension? Ice, rest, relaxation and sleep often helps to get rid of these.
Another big help for me is heat. I don't really like using heating pads on my neck and shoulder as they don't seem to lie the I need them to in order to get the most relief. I'm also not too fond of the conventional heating pads, as I have had patients inadvertently get hurt by them.
I have seen some great companies come up with safer and more practical heating pads in recent years. I still prefer to make my own.
What I have found that works best for me, are those rice bag heating pads that you heat in the microwave. These are great for many different areas of your body; they conform well around your neck, shoulders lower back etc. I love that you have full control over the size, thickness, density and heat level. Best thing of all, it's super cheap and you can customize it as you see fit.
If you don't already have one these rice bag heating pads or don't know how to make them, I'd like to share my simple way to make a DIY Homemade Heating Pad.
DIY Homemade Heating Pad
Things You're Going to Need:
Uncooked White Rice – Do not use brown rice as it will break up and get mushy after it's microwaved (the composition of it is different from white rice)
Thick Sock of your choice – men's tube type sock, sports socks, any type of sock that is of good thickness and the length you desire
New Hair rubber band with no metal inside
Ribbon – Optional
Small plastic cup – one that is thinner so it can be somewhat bent
Let's Put it Together:
You want a pretty thick sock so the rice doesn't seep through the fabric. If you are like me you'll be using this over and over again so you'll want it to last. I choose long thick soccer socks (unused) I love when the heating pad is on the longer side so when it's draped over my shoulders I can have it kind of hit my collarbone area. This sure does bring a lot of relief to my sore/aching neck and shoulders.
Take your sock and open it up and start dumping the rice in with your flexible cup. I use a small cup that fits in so I can do it pretty fast without getting it everywhere. Fill up the sock all the way leaving about an 1 1/2 to 2′ from the top. How firm you make it is completely up to you. When pouring in the rice you can pack it in or you can add less rice. This all depends on how heavy you want your heating pad and how flexible you want it.
Once you have it as full as you want wrap the hair rubber band around as many times as it can go. Make sure it's tight enough where the rice isn't gonna come out, those grains are tiny.
Finish off with ribbon around the rubber band if you want.
How to Use your DIY Homemade Heating Pad:
When you need to use your heating pad all you need to do is place it on a microwave safe dish. Heat it up; starting at about 1 minute 30 seconds and increase or decrease from there until it's at the desired heat level you want and can tolerate. Each microwave is different and each person's tolerance of temperature is different. Be careful and use pot holders if you need to. Remember different areas of your body are more sensitive to heat so test carefully.
You can use it on your neck, upper and lower back, shoulders, arms, legs, ankles, feet , just about anywhere.
Don't forget as with anything microwaved item, there can be areas that get hotter than others. Use with caution; especially with children and elderly.
You can always add or take out rice once you see how full you want it. If you plan on putting a few stitches in to keep it closed be sure to test it out and make sure the thickness is how you want it.
I love using the uncooked rice because it seems to hold the heat for a pretty long time.
I know there are lots of you out there that are crafty and can even make some DIY homemade heating pad covers for these. I've seen some pretty creative covers at craft fairs.
What tricks do you use to help with those headaches or tension aches and pains? Have you made a DIY homemade heating pad before?
Have you checked out my Giveaway Page to see all the latest giveaways?
I originally shared this project with the readers over at Sew Crafty Angel.
Close-Up Flowers in Watercolor (w/ Nan Carey)
Cooking Essentials: All About Pork (w/ Rick Rodgers)
Building Your Business: Beautiful Cake Photography (w/ Carrie Sellman)


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Wednesday, January 26, 2011
God With Us
This may seem unimpressive to us, who as 21st century Christians already assume that, but to an ancient Hebrew this was a pretty big deal. Gods didn't go with their people; people went to their Gods. The ancient world had holy places: holy mountains, holy temples, holy trees, etc. If a person wanted to be in the presence of the god or goddess, the person needed to find the place that god or goddess was and then go worship there. On top of that, the ancient world was largely pantheistic. In other words, the god or goddess and natural material were the same thing. The god or goddess was the thing. For example, when Moses saw the burning bush, a pantheistic Canaanite would have assumed the bush was a manifestation of the god. The bush would have been god, not a god speaking from a bush. That's why idols were so prevalent in the ancient world. By making an image in the form of a god or goddess, that idol became the god or goddess. I'm not sure how to make that idea clearer, but the point is the Hebrew God was a very different animal. The Tabernacle was not God, it was God's tent, just like the tents the Hebrews were living in. The Ark was not God, it was God's throne, the ultimate seat of justice, much like the throne Moses sat in when he judged over Israel. The God of Moses, Abraham, Isaac, and Jacob was not a god merged with a place or thing, but a dynamic God who had chosen to travel with this people. I would imagine from what the Hebrews were used to, that would have scared the bugeebees out of them. At the same time, it would have been a life changing, world re-framing event. The Holy One of Israel was in their midst.
Stopping point: Exodus 27
Labels: Ark of Covenant, Exodus, pantheism, Tabernacle
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- 4 beds
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- Half Baths: 1
- Square Feet: 2,408 sqft
- Year Built: 2006
- Lot Size: 2.006 acres
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Great Custom 1st floor master colonial situated on an amazing FENCED 2 acre lot with fabulous detail inside and out with a deep set back from the country style street near highways and shopping with energy savings from the GEOTHERMAL heating and cooling, Gourmet granite kitchen, Light filled rooms throughout! Newer winding paved drive will lead you to this attractive premium exterior with extra parking pad. Raised deck overlooks this ONE OF A KIND FULLY FENCED YARD! Enter to a dramatic spindled staircase and soaring ceilings in the 2 story great room that includes floor to ceiling windows for that added touch. Kitchen has been recently updated to include GRANITE COUNTERS, custom tiled backsplash and includes appliances...Warm and inviting dinette along with a bayed out formal dining room that is open to the foyer. Large 1st floor laundry. The 1st floor owner's suite will pamper you with huge closet and private glamour owner's bath! 3 additional bedrooms up with a lookdown to the great room. Spacious open basement that has endless possibilities. Country style large lot living at its best...Call today for your private showing. YOU WILL LOVE IT!
- Site Link
- Property Subtype Single Family Detached
Listing Agent: Christopher Frederick
Courtesy of: Coldwell Banker Hunter Realty
Office Phone: 4407778500
Updated: 21st August, 2017 12:13 PM.
13377 Quarry Rd Oberlin, OH


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Posted Under: Life & Time Management Hacks, Simple Things You can Do:
July 26, 2015
habits, scheduling tasks, software
Comments: 0
Windows Users: Scheduling Tasks With Windows
For a long time, I used a third party app to schedule tasks, especially repetitive tasks. Then one day, the app I used stopped working and I needed tech support. The company never responded (stupid - they lost a customer).
So, as I started looking for a third party replacement, I remembered that supposedly there was a way for Windows to be set up to do repetitive tasks. It turns out my memory was spot on.
And it's so simple that you can apply this tip - today!
The Windows Task Scheduler that is one of the System Admin Tools under the Windows Control Panel.
Setting up a task is simple - just use the Basic Task Wizard. I created some detailed instructions should you need them.
One nuance to tell you about. If you want the Action of the task to include going to a website destination, you need to enter two pieces of information.
1 - The browser to be used. Click Navigation, and locate your browser's .EXE file.
2 - In the Additional Arguments field enter the URL.
Otherwise, the program is very straightforward.
Play with it a few times, and you too can be a Task Scheduling pro.
P.S. - Did you find this information of use? Will you put the information you got here to use - today?
Yes? Then You Are Invited: Get your complimentary subscription to my ezine, Better Ways - Today and get notified when I create a new post about lots of different things that you can apply - today - or at least quickly.
Our journeys begin with initial steps and continue with more steps. Sometimes you take a step that accelerates your progress, and sometimes, these steps come from ideas you receive along the way. Give Better Ways - Today a try and see if you don't accelerate just a little bit more (perhaps a lot more).
I also have a free gift for new and current subscribers called, ShatterTime. that talks about how we waste our time, the monetary cost of it and some simple ideas about what to do about it. Simply provide a working email address.


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Wim Boswinkel
»The novel is an interesting amalgam of genres, drifting in and out of allegory, satire, romance and myth. It will not be well received by the bigots and the hypocrites in our midst and that is to its great credit.«
John Eppel
Johan comes to Africa to manage a tea plantation. He meets Erina, and his life changes forever. The story takes a leap into the unknown, cleverly blending an African setting with the fantastic premise at its core: the arrival of a black female Christ-figure. The use of AIDS as a weapon to effect the ultimate defeat of Satan adds a powerful and provocative dimension.
Erina won Best First Book at the Zimbabwe Book Publishers Association awards


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// Unwanted Fat
Workout to Beat the Winter Blues
Having the winter blues? Join the club. If you're anything like me, then you see red whenever you see snow. For those of us who were hoping that maybe winter would meet an early demise this year, this week marks a very disheartening part of the calendar. Yes, we are still in the month of February, and we shouldn't expect the cold to have disappeared yet – but somehow we wish it would anyway.
For many of us, we dream of the days when the weather will warm up and we'll be able to get back to our regular exercise routines. But, I'm here to tell you that cold weather is no excuse to give up on your dedication to fitness. Sure the cold doesn't make it easy to get exercise outdoors. But it doesn't make it impossible, either! Believe it or not, you do have options.
On SparkPeople.com, certified personal trainer, Jennipher Walters explains that there are many ways to stay active throughout the long winter season. It begins with a positive attitude, she says. In other words, for those of us that take one look outside and allow for the wintery atmosphere to get us down, it's time we snap out of it! The following are some ways to make the most out of your winter workouts.
Change your mind. To reiterate, it seems as if some of us are in need of an attitude adjustment. Walters reminds us that winter is a time of year when we should take the time to sit down and revisit our goals. We often keep ourselves so busy that we neglect some of the important things in our lives – our health being one of them. Take time to reflect and plan out the goals you wish to accomplish over the next several months.
Go out and play! Who says that having fun in the snow is only for the kids? Perhaps, the best way to get your winter workout in is to simply play outside. This calorie-burning activity won't even seem like a workout thanks to all of the fun you'll be having. Take the kids out and have a snowball fight (just be careful!). Walters notes that you can burn more than 300 calories by doing so. Not enough snow? Perhaps, it's time to go ice skating!
Get creative at home. Naturally, the indoors are more appealing during the cold winter months. So why not turn your winter home into a workout home? Pop in a workout DVD and turn your living room into a makeshift gym. Walters also suggests that you invest in some fitness equipment. It will help for the comfort of your own home to be both a convenient and fun location to shed some extra pounds during the winter.
Try something new. For those that prefer the indoors during winter – remember, you are not alone! – the season marks the perfect time for you to sign up for a new class or indoor sports league. There are plenty out there to choose from including "indoor volleyball, a dodgeball league, a bootcamp class or even tennis lessons at a local indoor racquet club" suggests Walters.
Eat right. Of course, cold weather is no excuse for you to forget about the importance of nutrition. You're always doing yourself a favour when you remember to limit your sweets and eat plenty of healthy foods. Walters reminds us that in-season produce such as oranges, kale and chestnuts make great healthy snacks. Try to embrace winter while you can as it won't last forever. Thankfully!


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Clients Value Phil Van Hooser's Leadership Keynotes and Training
Leadership keynotes and training presentations delivered by Phil Van Hooser have received overwhelming positive feedback from the hundreds of clients he's worked with. In fact, they value the results so much, that 2/3 of Phil's clients ask him to work with their leaders more than once.
- Alliance Resource Partners, LP
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- Eli Lilly
- ExxonMobil
- Helena Chemical Company
- KeyBank
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- Mutual of Omaha
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Phil Van Hooser is the best received speaker in the ten years of our Supervisor Leadership training. - Westar Energy
There is so much "noise" about leadership these days. But your material is different because it is hands on and based on your experience across various platforms. Thank you for all you do and your encouragement. - Doug C.
The message delivered hit not only home -- but a HOME RUN! - APPA
Phil Van Hooser has helped my team immensely. He has adapted his style to every situation we have utilized him for and the results are just outstanding. Phil has become a part of our team and not just another consultant. - Helena Chemical Company
The feedback we received was the best we have ever received from our group. They walked away with so much good information to help them on their personal leadership development. Everyone thought he was a 'rock star!' They wanted more… everyone wanted him to speak all day. - F. F. Properties L.P.
…this last session was the Grand Slam Home Run, the best of the best so far. - Blue Bell Creameries
You were definitely a hit with the entire audience! My guess is you had close to a 100% approval rating. I have never heard that much positive feedback or excitement about an outside presenter at Lilly. The areas that you focused your presentation on were spot on... - Lilly Research Laboratories


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With the emergence of YouTube, inexpensive webcams and widespread broadband Internet access, innovative forms of e-recruitment have become a norm. Unlike the ".docx" version, video resumes—or visumes—allow you to demonstrate interpersonal, creative and technical skills to your employer, while showcasing your personality in a matter of minutes.
A behind the scenes look
"Video resumes are an excellent way to get companies to sit up and take notice—if you do it right," says Jyotika Dhawan, co-founder and managing director of Helix HR, New Delhi. "They give a more personal view of you than a paper resume. They allow employers to see how you present yourself, and whether you would fit into the company culture. If you make the right impression, your video resume could get you to the interview stage quicker."
For all that, a trend that has thousands of takers the world over still has a long way to go in India. In 2007, when Monster.com introduced video resumes in India as a new feature on the site, the e-recruiting market was excited and hopeful. However, the lack of specialized professionals to aid candidates' efforts, combined with their inability to shoot and edit their own videos, made it a damp squib. "The trend really hasn't taken off in India partly because it isn't convenient yet for employers to 'search' a video resume among thousands of applications," says Sanjeev Bikhchandani, founder and CEO of? Naukri.com, and a Business of Life columnist. "But more importantly, because the real challenge lies in doing a good job of it—and that's not as easy as it seems."
However, with the freeze on recruitments finally easing up, what better time to focus on rebranding your professional image? Just remember that visumes must walk a delicate line, maintaining a professional approach while giving the employer a personality-filled, attention-grabbing glimpse of you. Here's how to make yours a job-winner:
Look the part
"If you're applying for a position in a formal, traditional company or industry such as banking your video resume may not be the place to showcase your Zumba dancing skills or your passion for stand-up comedy," says Dhawan. "However, if you're looking for a more creative field such as advertising, you may be able to get away with a little more fun element." Know your sector and highlight your job-related skills. Your visume is your chance to make a compelling first impression—show employers you can fit in.
Run a quality check before you 'publish'
It may be a good idea to approach a professional corporate film-maker to produce your video resume. However, if you're planning to make it on your own, make sure you give some thought to the technical aspects of the recording. Bad lighting, inappropriate settings, poor editing and audio glitches can completely ruin your chances. "The impression you make will depend a whole lot on the overall look and feel of your video," says Neeraj Ram, a New Delhi-based professional film-maker and founder of Film Wallahs. "Keep it clean and simple to make sure the employer has no distractions and can concentrate on you. Be patient and ready to review the clips several times over during the shooting process to avoid any blunders." Ask friends, family and trusted colleagues to review it too.
Prepare to face the camera
Unless you've been on camera before, it may take you some time to relax in front of the lens. "Take the time to pen a script before you get down to filming, so you know exactly what you're going to say and can focus on how you're going to say it," advises Ram. "Be relaxed, and avoid big, sweeping or repetitive gestures—but don't be afraid to let your personality come through."
Make it the prologue to your pitch
Remember, however, that though a video resume is a fantastic personal marketing tool, it does not replace your text resume. Look on it as a teaser or a sales pitch to get recruiters excited about you and leave the details of your job experience to your CV. Also, remember that hiring managers potentially have a pile of applications on their desks and do not have time for a sitcom. To avoid putting them off, keep your message short and relevant—no longer than 1-3 minutes. A visume made by a professional film-maker can cost upwards of Rs15,000 and will need a minimum of two full days of shooting and couple of days for editing.
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