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Conserved glycine 232 in the ligand channel of ba3 cytochrome oxidase from Thermus thermophilus.

Biochemistry (2014-06-18)
William McDonald, Chie Funatogawa, Yang Li, Ying Chen, Istvan Szundi, James A Fee, C David Stout, Olöf Einarsdóttir
ABSTRACT

Knowing how the protein environment modulates ligand pathways and redox centers in the respiratory heme-copper oxidases is fundamental for understanding the relationship between the structure and function of these enzymes. In this study, we investigated the reactions of O2 and NO with the fully reduced G232V mutant of ba3 cytochrome c oxidase from Thermus thermophilus (Tt ba3) in which a conserved glycine residue in the O2 channel of the enzyme was replaced with a bulkier valine residue. Previous studies of the homologous mutant of Rhodobacter sphaeroides aa3 cytochrome c oxidase suggested that the valine completely blocked the access of O2 to the active site [Salomonsson, L., et al. (2004) Proc. Natl. Acad. Sci. U.S.A. 101, 11617-11621]. Using photolabile O2 and NO carriers, we find by using time-resolved optical absorption spectroscopy that the rates of O2 and NO binding are not significantly affected in the Tt ba3 G232V mutant. Classical molecular dynamics simulations of diffusion of O2 to the active site in the wild-type enzyme and G232V mutant show that the insertion of the larger valine residue in place of the glycine appears to open up other O2 and NO exit/entrance pathways that allow these ligands unhindered access to the active site, thus compensating for the larger valine residue.

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