The same hydration and dehydration experiments were repeated on bulk samples of microcrystalline Co-MAHF-9 A8Q-hydrated to verify the homogeneity of the transformation, and PXRD was used to track the unit cell changes (Figure ). Pawley refinement (Figure ) of Co-MAHF-9 A8Q-hydrated gave unit cell parameters that agree very closely with those derived from SC-XRD data (Table ). Decreasing the water content to 50% H2O/50% MeCN did not alter the powder pattern nor the color of the solid. However, removing all water yielded a purple solid, and Pawley refinement shows a unit cell characteristic of Co-MAHF-9 A8Q-dehydrated. Co-MAHF-9 A8Q-dehydrated can be fully rehydrated by soaking in 100% water, and PXRD verifies that the material structure has returned to that of Co-MAHF-9 A8Q-hydrated. The coupling of metal coordination changes with helix movements suggests the possibility of allostery, where local structural changes influence neighboring sites, which could lead to highly coherent framework phase transitions due to the positive cooperativity. To test this hypothesis, diffuse-reflectance UV-vis spectroscopy was used to monitor the coordination geometry of Co(II) as water was titrated into a suspension of Co MAHF-9 A8Q-dehydrated in acetonitrile (Figure ). The electronic adsorption spectrum of tetrahedral Co(II) persisted until 25% H2O was reached, at which point, a sharp transition to octahedral Co(II) occurred. The sigmoidal titration curve is a signature of positive cooperativity. As some Co(II) sites begin to change geometry, the helices coordinated to them also move, causing the adjacent helices packed against the sites to also shift. These movements trigger conformational changes to distant Co(II) sites. The A8Q mutation is responsible for generating the complex phenomenon of positive cooperativity. As shown in Figure , the parent framework, Co-MAHF-9, does not show any significant spectral changes in response to water content in its environment (Figure ). One possible reason that the A8Q mutation results in cooperativity and flexible coordination chemistry is that Gln8 can make H-bonds with Co-OH2 moieties or water molecules in the case of Co- MAHF-9 A8Q-hydrated or with Co-His units in the case of Co- MAHF-9 A8Q-dehydrated. However, Co-MAHF-9 has a hydrophobic pore that prefers conformations that bury Ala8. It is remarkable that a single mutation can switch on stimuli-responsive cooperativity, and it underscores the evolvability of peptide-based materials. Structure analyses. All framework variants possess alternating metal-coordinated and hydrophobic layers of its parent structure, Co-MAHF-9, demonstrating the ability of the MAHF-9 family to vary while inheriting some traits from its predecessors. All peptides fold into ɑ -helices with no significant distortions (Figure ). However, the spread of ɸ/ψ angles indicates that even an ideal ɑhelix is semi-flexible, and peptide components can slightly adjust their structure to maximize favorable interactions. Additionally, the sidechains of Glu4 and His7 can freely rotate about the χ torsions and bind to metal ion in various modes; Glu can bind as κ 1 or κ 2 , with the former making either cis or trans Cɣ-Cδ-O-M linkages, and His can coordinate with either Nε or Nδ. The semi-flexibility of the peptide is likely advantageous for framework formation as it allows the system to find assembly solutions that satisfy both metal coordination and packing of mutated residues. The versatility of this helix building block is evidenced by the fact that 16 of the canonical amino acid mutations and several transition metals readily produce singlecrystalline frameworks, with all the rest producing microcrystalline powders. There are multiple degrees of freedom for the assembly quaternary structure to adjust in response to the A8X mutations, as helix chains can move in vicinity, rotate, flip, and slide. Since the metal ion is bound to at least two different helices, the relative orientation of the helices directly impacts the metal coordination environment. The exact packing of the helices is highly specific to the size, shape, and electronic properties of residue 8, making it difficult to draw broad trends. For example, Co-MAHF-9 A8S and Co-MAHF-9 A8T assemble to form significantly different Co(II) frameworks with minimal structural differences (a methyl group in this case). On the contrary, Co-MAHF-9 and the Co-MAHF-9 A8F form very similar structures despite the drastic size difference between Ala and Phe. However, these structural differences can be rationalized on a caseby-case basis, and they are explained by the specific steric, hydrophobic, and H-bonding profile of the helix, as well as the coordination preferences of the metal ion. The ability for residue 8 to H-bond strongly influences the metal coordination sphere. With Co(II), all amino acids possessing H-bonding sidechains (except Trp8 and Arg8) form a secondary sphere interaction with either a metalbound water ligand or Glu4/His7 via H-bonds. Transition metals with stronger coordination preferences can override the H-bonding influences, as seen in the frameworks derived from the A8S mutant with Mn(II), Fe(II), Cu(II), and Zn(II). Overall, the ɑ -helix is shown to be a stable and robust building block for framework assembly, with the majority of framework changes occurring at the quaternary structure level due to non-covalent interactions. Furthermore, the vast properties of canonical amino acids enable the production of distinct primary and secondary metal coordination spheres by a single-point mutation within the sequence. Point mutations of MAHF-9 generate several types and permutations of coordination spheres, spanning 4, 5, and 6 coordinate numbers, as well as various coordination geometries and stereoisomers. Among the 4-coordinate metal sites were tetrahedral, distorted tetrahedral, see-saw, trigonal pyramidal, and square planar geometries. Most of the 4-coordinate centers have 2His-2carboxylate ligand sets, but Co-MAHF-9 A8D and Co-MAHF A8C possess 1His-3carboxylate and 2His-2Cys compositions, respectively. For 5-coordinate centers, only examples with square pyramidal geometry were found (Co-MAHF-9 A8Q, Co-MAHF-9 A8Q-hydrated, and Mn- MAHF-9 A8S), though they each possess unique ligand sets varying in the amounts of His, Glu, and water. There are several types with 6-coordinate metal sites in the MAHF-9 series. Both octahedral and the rarer trigonal prismatic (Co-MAHF-9 A8M) geometries were found. For octahedral sites, the peptide framework may act as a bito hexadentate ligand, and within the tridentate subset, 2His-1Glu (Co-MAHF-9 A8T, Co-MAHF-9 A8N) and 1His-2Glu (Co-MAHF-9 A8W) compositions are seen, which can be further classified as facial, trans-meridional, or cis-meridional isomers.