Disruption of a hydrogen bond network in human versus spider monkey cytochrome c affects heme crevice stability

• Published in: Journal of inorganic biochemistry Vol. 158; pp. 62 - 69
• Main Authors: Goldes, Matthew E., Jeakins-Cooley, Margaret E., McClelland, Levi J., Mou, Tung-Chung, Bowler, Bruce E.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01-05-2016
• Subjects: Alkaline conformational transition; Amino Acid Substitution; Animals; Atelinae; Circular Dichroism; Crystallography, X-Ray; Cytochromes c - chemistry; Cytochromes c - metabolism; Global stability; Heme - chemistry; Heme - metabolism; Heme crevice stability; Humans; Hydrogen bond network; Hydrogen Bonding; Hydrogen-Ion Concentration; Kinetics; Protein Folding; Protein Stability; Spider monkey cytochrome c; Spider monkey cytochrome c; Hydrogen bond network; Heme crevice stability; Global stability; Alkaline conformational transition
• ISSN: 0162-0134; 1873-3344
• DOI: 10.1016/j.jinorgbio.2015.12.025

The hypothesis that the recent rapid evolution of primate cytochromes c, which primarily involves residues in the least stable Ω-loop (Ω-loop C, residues 40–57), stabilizes the heme crevice of cytochrome c relative to other mammals, is tested. To accomplish this goal, we have compared the properties of human and spider monkey cytochrome c and a set of four variants produced in the process of converting human cytochrome c into spider monkey cytochrome c. The global stability of all variants has been measured by guanidine hydrochloride denaturation. The stability of the heme crevice has been assessed with the alkaline conformational transition. Structural insight into the effects of the five amino acid substitutions needed to convert human cytochrome c into spider monkey cytochrome c is provided by a 1.15Å resolution structure of spider monkey cytochrome c. The global stability for all variants is near 9.0kcal/mol at 25°C and pH7, which is higher than that observed for other mammalian cytochromes c. The heme crevice stability is more sensitive to the substitutions required to produce spider monkey cytochrome c with decreases of up to 0.5 units in the apparent pKa of the alkaline conformational transition relative to human cytochrome c. The structure of spider monkey cytochrome c indicates that the Y46F substitution destabilizes the heme crevice by disrupting an extensive hydrogen bond network that connects three surface loops including Ω-loop D (residues 70–85), which contains the Met80 heme ligand.
Destabilization of the heme crevice during the rapid evolution of primate cytochromes c is primarily due to a Y46F substitution, which disrupts a hydrogen bond network that knits together three surface Ω-loops. [Display omitted]
•The rapid evolution of primate cytochromes c affects heme crevice stability.•The heme crevice stability of spider monkey and human cytochrome c is compared.•The Y46F substitution is the primary destabilizer of the heme crevice.•A 1.15Å resolution structure of spider monkey cytochrome c is presented.•The Y46F substitution disrupts a hydrogen bond network connecting 3 surface Ω-loops.