title: Thermal stability of hydrophobic heme pocket variants of oxidized cytochrome c.
abstract: Microcalorimetry has been used to measure the stabilities of mutational variants of yeast iso-1 cytochrome c in which F82 and L85 have been replaced by other hydrophobic amino acids. Specifically, F82 has been replaced by Y and L85 by A. The double mutant F82Y,L85A iso-1 has also been studied, and the mutational perturbations are compared to those for the two single mutants, F82Y iso-1 and L85A iso-1. Results are interpreted in terms of known crystallographic structures. The data show that (1) the destabilization of the mutant proteins is similar in magnitude to that which is theoretically predicted by the more obvious mutation-induced structural effects; (2) the free energy of destabilization of the double mutant, F82Y,L85A iso-1, is less than the sum of those of the two single mutants, almost certainly because, in the double mutant, the -OH group of Y82 is able to protrude into the cavity formed by the L85A substitution. The more favorable structural accommodation of the new -OH group in the double mutant leads to additional stability through (1) further decreases in the volumes of internal cavities and (2) formation of an extra protein-protein hydrogen bond.
authors.affiliation: Department of Biochemistry, University of Texas Health Science Center, San Antonio 78229-3900, USA.
authors.forename: J R
authors.initials: JR
authors.lastname: Liggins
authors.forename: T P
authors.initials: TP
authors.lastname: Lo
authors.forename: G D
authors.initials: GD
authors.lastname: Brayer
authors.forename: B T
authors.initials: BT
authors.lastname: Nall
mesh_terms: D019943:Amino Acid Substitution; D002151:Calorimetry; D018360:Crystallography, X-Ray; D003574:Cytochrome c Group / Q000737:chemistry* / Q000235:genetics; D006418:Heme / Q000737:chemistry* / Q000235:genetics; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011489:Protein Denaturation; D017510:Protein Folding; D012441:Saccharomyces cerevisiae / Q000737:chemistry; D013816:Thermodynamics
chemical_list: D003574:Cytochrome c Group; D007527:Isoenzymes; D006418:Heme