title: The role of position a in determining the stability and oligomerization state of alpha-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.
authors.affiliation: Department of Biochemistry and the Medical Research Council Group in Protein Structure and Function, University of Alberta, Edmonton, Canada.
authors.forename: K
authors.initials: K
authors.lastname: Wagschal
authors.forename: B
authors.initials: B
authors.lastname: Tripet
authors.forename: P
authors.initials: P
authors.lastname: Lavigne
authors.forename: C
authors.initials: C
authors.lastname: Mant
authors.forename: R S
authors.initials: RS
authors.lastname: Hodges
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D004220:Disulfides; D004355:Drug Stability; D019791:Guanidine; D046911:Macromolecular Substances; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry*; D012044:Regression Analysis; D013816:Thermodynamics; D014508:Urea
chemical_list: D004220:Disulfides; D046911:Macromolecular Substances; D010455:Peptides; D011506:Proteins; D014508:Urea; D019791:Guanidine