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Naturally Occurring Disease-Related Mutations in the 40–57 Ω-Loop of Human Cytochrome c Control Triggering of the Alkaline Isomerization

Deacon, Oliver M and Svistunenko, Dimitri A and Moore, Geoffrey R and Wilson, Michael T and Worrall, Jonathan AR (2018) 'Naturally Occurring Disease-Related Mutations in the 40–57 Ω-Loop of Human Cytochrome c Control Triggering of the Alkaline Isomerization.' Biochemistry, 57 (29). pp. 4276-4288. ISSN 0006-2960

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American Chemical Society. Naturally occurring mutations found in one of the two ω-loop substructures in human cytochrome c are associated with low blood platelet count (thrombocytopenia). Both ω-loops participate in the formation of conformers associated with cytochrome c peroxidase activity and apoptotic function. At alkaline pH values, the Met80 ligand to the ferric heme iron dissociates, and a lysine residue in the 71-85 ω-loop coordinates to the iron. The alkaline isomerization has been the focus of extensive kinetic studies, and it is established that a deprotonation triggers the release of the Met80 ligand (pKtrigger). A second deprotonation stabilizes a pentacoordinate heme form (pKa2). In this study, site-directed variants at the 41 and 48 positions in the 40-57 ω-loop and at the 81 and 83 positions in the 71-85 ω-loop reveal that conformational transitions in the 71-85 ω-loop, leading to the alkaline or peroxidatic conformers, are controlled by the 40-57 ω-loop. We find that the variants causing thrombocytopenia, G41S and Y48H, lower the pKtriggerand increase pKa2. Our results are presented in a mechanistic framework, depicted by a cube, that accounts for the pH dependencies of the equilibrium and kinetic parameters governing the alkaline transition of the native protein and ω-loop variants. The data are most consistent with the trigger for Met80 replacement by a lysine being a deprotonation within a hydrogen bonded unit that links the two ω-loops rather than an individual group. Such a proposal aligns with the entatic contribution made by the same unit in controlling the Met80-Fe(III) bond strength.

Item Type: Article
Uncontrolled Keywords: Humans; Thrombocytopenia; Alkalies; Cytochromes c; Peroxidase; Protein Conformation; Protein Denaturation; Kinetics; Point Mutation; Isomerism; Hydrogen-Ion Concentration; Models, Molecular; Protein Stability
Subjects: Q Science > QD Chemistry
Q Science > QH Natural history > QH301 Biology
Divisions: Faculty of Science and Health
Faculty of Science and Health > Life Sciences, School of
SWORD Depositor: Elements
Depositing User: Elements
Date Deposited: 20 Sep 2018 14:52
Last Modified: 06 Jan 2022 13:51

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