the cause of various diseases involves the human leukocyte antigen ( hla ) system which is the human version of the major histocompatibility complex . the hla genes involved in the immune response fall into two classes , i and ii , which are structurally and functionally different . the association between specific hla alleles and adverse drug reactions which frequently cause significant morbidity and mortality for patients is also widely known . a reliable three - dimensional ( 3d ) structure of the particular hla allele responsible for the specific event is essential to understand the underlying molecular mechanism in order to develop effective therapeutic agents or / and countermeasures . since the pioneering work by wiley et al . the crystal structures have shown that the peptide - binding groove of an hla molecule consists of two parts , a floor and two walls . although this canonical topology is highly conserved among different alleles , certain structural differences exist depending on the alleles . therefore , the 3d structure of a particular hla molecule is required for hla studies . the hla genes are the most polymorphic in the human genome and there are a large number of allelic variations . in the case of alleles belonging to the isotype a of class i hla , the number of independent alleles deposited in the imgt / hla database   is 1,372 as of 27 july 2013 ( version 3.13.1 ) . on the other hand , the number of the corresponding nonredundant alleles whose 3d structures are deposited in protein data bank   as of 24 april 2013 is just nine . this shows that a huge gap exists between known allele sequences and available 3d structures . therefore , it is reasonable to assume that experimental structures for all the possible hla alleles will not be available in the near future . in the absence of experimental structures , in silico homology modeling can provide a viable alternative to generate reasonably accurate models of the allele of interest . homology modeling is a methodology to predict protein structure based on the general observation that proteins with similar sequences have similar structures . the accuracy of homology models compared to the actual experimental structure is generally judged by c atomic pair root - mean - square deviation ( rmsd ) . depending on the degree of sequence identity or similarity and the quality of the alignment , the rmsd can be up to ca . 1 - 2   . homology modeling package specialized in hla molecules is not available until now as far as we know . the purpose of this study is to create an automated hla modeling application suitable for hla studies at the molecular level . the application named hla - modeler was coded by the use of scientific vector language ( svl ) implemented in moe . all of the crystal structures of the hla molecules deposited in the pdb were retrieved and a local hla structural database named hla-3ddb was compiled on april 24th , 2013 . the proper allele names were obtained from imgt / hla . the template structure which is most homologous to the query sequence the optimization of the models is carried out by use of the pfrosst force field [ 10 , 11 ] considering the implicit solvent model . the best structure in terms of the free energy of hydration calculated based on generalized born / volume integral implicit solvent model   is selected . in the final optimization of the structure , nonhydrogen atoms are tethered . the antigenic peptide - binding site is primarily used in hla - modeler in order to best use the 3d characteristics of the essential site of the template structure . a flow chart of hla - modeler specific binding of antigenic peptides to a particular hla molecule is a central problem for most of the hla studies . hla - modeler can construct the homology model of hla - peptide complex based on the supplied peptide sequence . the svl code of hla - modeler is available from ryoka systems inc . on request . it is of interest to validate the reproducibility of homology models routinely built by hla - modeler . for this purpose , the leave - one - out cross - validation was undertaken . in the current hla-3ddb , there are 41 and 27 crystal structures of nonredundant hla molecules belonging to classes i and ii , respectively . for each amino acid sequence in hla-3ddb , homology models were constructed using all structures belonging to the same class except the identical structure as template structures . the results for classes i and ii are illustrated in figures 2(a ) and 2(b ) , respectively . the rmsds do not depend on the sequence identity and are generally lesser than 1.1  . the only exception is the structure of hla - dr1 ( pdb i d : 4gbx ) modeled based on the template structure of hla - dr52c ( 3c5j ) . since the structure of 4gbx is the heterodimer between hla - dr1 and hla - dm , the structure of the hla - dr1 molecule is deformed due to the interaction with hla - dm . if all domains are used for homology modeling , rmsds are up to 3.0 and 2.7   in classes i and ii , respectively . it indicates that the homology modeling strategy adopted in hla - modeler is suitable to construct the structure of antigenic peptide - binding site . in the first case , a homology model was built using the sequence of the hla - c04:01 molecule ( pdb i d : 1qqd ) as a query sequence , and the structure of the hla - b44:02 molecule ( pdb i d : 1m60 ) is selected as a template structure . the nonhydrogen atoms of the 1 and 2 domains of the homology model and the crystal structure are superimposed in figure 3 . it is considered that such degree of discrepancy as shown in figure 3 may be small enough for most qualitative analysis such as estimation of amino acid residues which should possibly bind to antigenic peptides . even in the cases where sequence identity is low , reasonably accurate models can be constructed as illustrated in figure 2 . however , if it is necessary to predict the conformations of amino acid residues at the peptide - binding site as accurate as possible , it is better to use the template structure with higher sequence identity . in the second case of homology modeling of the hla - a02:01 molecule ( pdb i d : 1lp9 ) based on the template structure of a mutant molecule of hla - a02:01 ( 2uwe ) , the sequence identity is very high ( 99.2% ) . only one residue at the antigenic peptide - binding site is different , that is , ala and thr in 2uwe and 1lp9 , respectively . the rmsd of nonhydrogen atoms of the peptide - binding sites is 0.23   . the position of the relevant thr residue in the homology model is almost identical to that in the crystal structure . in summary , we can construct reasonably reliable 3d structures of class i hla molecules by hla - modeler . hori et al.found that the disease is significantly associated with hla - dp5 and identified an immunodominant peptide . understanding the interactions between the peptide and the hla molecule at the molecular level will greatly help to find therapeutic strategies against this disease . however , the 3d structure has not been disclosed so far . by use of hla - modeler , we have built the homology model of the hla - dp5 molecule with the minimum immunodominant peptide . the main chains of the antigenic peptides bound to the hla molecules take highly conserved extended structures . in particular , the main chain structure of eight residues shown in figure 5 is conserved . if the main chain atoms of the corresponding eight residues in 28 independent peptides bound to the class ii hla molecules in the crystal structures are superimposed , the median rmsd for each residue ranges from 0.26 to 0.50 . it indicates that the main chain conformations of the peptides bound to class ii hla are conserved in this particular region . the eight residues involving p1 , p4 , and p6 anchoring residues should play significant role in binding to the class ii hla molecules . by taking the structural conservation of the bound peptides into account , the structures of the complexes between the hla dp5 molecule and the nonapeptide with seven different alignments were optimized . the binding affinity of each complex was judged by a scoring function of gbvi / wsa_dg which is considered to express protein - ligand binding free energy . the complex structure with the minimum gbvi / wsa_dg value is shown in figure 6 . the two terminal k and f residues of the peptide protrude from the antigenic peptide - binding groove and point toward the t - cell receptor . the experimental data have demonstrated that these residues are essential for the interactions with t - cell receptor . since there are a large number of allelic variations , it is expected that the 3d structures of all the possible hla alleles will not be experimentally determined in the near future . under these circumstances , it is indispensable to use in silico methodology to predict the missing structures to promote hla studies . the present study has demonstrated that if we properly use a strategy to build up homology models based on the structurally conserved antigenic peptide - binding site , we can build the reliable 3d structure of the site which is essential for the functions . the 3d structures of particular hla molecules are useful to deepen our understanding of the molecular interactions between the hla molecules and specific antigenic peptides . the 3d models of hla molecules are also essential to disclose the molecular mechanisms of adverse drugs reactions closely related to specific hla molecules . moreover , the 3d models of the hla molecules associated with certain autoimmune diseases will contribute to the discovery of drugs which could suppress the autoimmune reactions . the automatic modeling system such as hla - modeler will be indispensable for extensive studies on these topics .
the three - dimensional ( 3d ) structures of human leukocyte antigen ( hla ) molecules are indispensable for the studies on the functions at molecular level . 
 we have developed a homology modeling system named hla - modeler specialized in the hla molecules . 
 segment matching algorithm is employed for modeling and the optimization of the model is carried out by use of the pfrosst force field considering the implicit solvent model . in order to efficiently construct the homology models , hla - modeler uses a local database of the 3d structures of hla molecules . 
 the structure of the antigenic peptide - binding site is important for the function and the 3d structure is highly conserved between various alleles . 
 hla - modeler optimizes the use of this structural motif . 
 the leave - one - out cross - validation using the crystal structures of class i and class ii hla molecules has demonstrated that the rmsds of nonhydrogen atoms of the sites between homology models and crystal structures are less than 1.0   in most cases . 
 the results have indicated that the 3d structures of the antigenic peptide - binding sites can be reproduced by hla - modeler at the level almost corresponding to the crystal structures .