The role of position a in determining the stability and oligomerization state of α‐helical coiled coils: 20 amino acid stability coefficients in the hydrophobic core of proteins

Abstract

We describe here a systematic investigation into the role of position a in the hydrophobic core of a model coiled-coil protein in determining coiled-coil stability and oligomerization state. We employed a model coiled coil that allowed the formation of an extended three-stranded trimeric oligomerization state for some of the analogs; however, due to the presence of a Cys-Gly-Gly linker, unfolding occurred from the same two-stranded monomeric oligomerization state for all of the analogs. Denaturation from a two-stranded state allowed us to measure the relative contribution of 20 different amino acid side chains to coiled-coil stability from chemical denaturation profiles. In addition, the relative hydrophobicity of the substituted amino acid side chains was assessed by reversed-phase high-performance liquid chromatography and found to correlate very highly (R = 0.95) with coiled-coil stability. We also determined the effect of position a in specifying the oligomerization state using ultracentrifugation as well as high-performance size-exclusion chromatography. We found that nine of the analogs populated one oligomerization state exclusively at peptide concentrations of 50 microM under benign buffer conditions. The Leu-, Tyr-, Gln-, and His-substituted analogs were found to be exclusively three-stranded trimers, while the Asn-, Lys-, Orn-, Arg-, and Trp-substituted analogs formed exclusively two-stranded monomers. Modeling results for the Leu-substituted analog showed that a three-stranded oligomerization state is preferred due to increased side-chain burial, while a two-stranded oligomerization state was observed for the Trp analog due to unfavorable cavity formation in the three-stranded state.

The Size and Conservation of a Coiled-coil Structure in the Ectodomain of Human BST-2/Tetherin Is Dispensable for Inhibition of HIV-1 Virion Release

“…In that study, alanine mutagenesis was well tolerated across much of the BST-2 ectodomain and only changes near the N and C termini of the ectodomain significantly affected BST-2 function (39). These results may not be surprising if one takes into consideration that alanine is the least destabilizing residue to add to the interaction surface of an oligomeric coiled-coil (55)(56)(57) and that replacing four consecutive residues at a time by alanine would not be expected to greatly disrupt the structure and interaction of the BST2 coiled-coil (58,59). Because of these limitations, previous alanine scanning mutagenesis did not recognize the importance of the central hinge region nor the relative unimportance of the size of the BST-2 ectodomain.…”

Preferred side-chain constellations at antiparallel coiled-coil interfaces

“…This limited set of rules has allowed considerable progress in the areas of coiled-coil prediction and design (1, 3), but our findings suggest that additional advances will be possible based on a more sophisticated treatment. Previously elucidated factors include lateral pairing of nonpolar side chains (9-16), which is driven by stereochemical complementarity, and lateral pairing of polar side chains (9)(10)(11)(12)(13)(14)(15)(16)33), which is driven by hydrogen bonding (at least for Asn-Asn pairs). In addition, pairing preferences are influenced by the electrostatic interactions (attractive or repulsive) between ionized side chains (34)(35)(36)(37)(38)(39).…”

“…The effects of sequence variation on coiled-coil stability have been widely studied (9)(10)(11)(12)(13)(14)(15)(16), but substantial gaps in our understanding remain. For example, the vast majority of prior efforts have focused on parallel coiled-coil homo dimers, but other coiled-coil states are seen in Nature.…”

Computational analysis of residue contributions to coiled‐coil topology

“…5 Thus, due to restrictions in packing geometry at a given heptad position, different residues will have different oligomeric propensities. 6,7 Recently, we used a molecular dynamics (MD) approach to dissect the oligomerization preferences of four coiled-coil sequences. 8 Our calculations reproduced some generally accepted rules such as the dimer specificity of LEU at d, the tetramer specificity of LEU at a, and the role of nonpolar side chains at positions e and g in specifying higher order structures.…”

“…17 A pattern of ILE at d with ALA at a creates a cavity in the interior of the parallel homodimer, while it generates a well packed interface in the antiparallel homodimer. 6 The effect of steric matching is more pronounced in the antiparallel dimer formed by a fragment of the C-terminal domain of the HIV-1 regulatory protein Vpr. The most prominent feature is a pair of TRP[a]/HIS[d] stacking interactions.…”