STRUCTURE OF THE NATIVE CYSTEINE-SULFENIC ACID REDOX CENTER OF STREPTOCOCCAL NADH PEROXIDASE. Joanne I. Yeh[[section]], Al Claiborne[[daggerdbl]], and Wim G.J. Hol[[section]], [[section]] Biomolecular Structure Center, Department of Biological Structure and Howard Hughes Medical Institute Box 357742, University of Washington, Seattle, Washington 98195-7742. [[daggerdbl]] Department of Biochemistry, Wake Forest University Medical Center, Winston-Salem, North Carolina 27157.
The flavoprotein NADH peroxidase (NPX) from Streptococcus faecalis represents one of two known flavin-dependent hydroperoxidases. Its flavin-linked function is chemically similar to flavoprotein disulfide reductases such as glutathione reductase. However, in NPX, an unusual, stabilized cysteine-sulfenic acid (Cys-SOH) is the redox center, in contrast to the redox active disulfide group of glutathione reductase. Indirect evidence for the existence of the Cys-SOH moeity resulted from metabolic labeling studies and FAB-mass spectrometric analysis of an active-site cysteinyl peptide (Poole and Claiborne, 1989). However, the initial x-ray crystal structure of the wild-type peroxidase refined at 2.16 Å resolution revealed that the Cys42 had been oxidized to the inactive sulfonic-acid form (Cys-SO3H) (Stehle et. al, 1991).
In order establish the existence of the Cys-SOH form of the NPX, the x-ray crystal structure of the peroxidase with its native Cys42-sulfenic acid redox center was determined. To obtain the native, unoxidized enzyme, a strategy combining reduced exposure to ambient oxygen and data collection at -160deg.C was used. The structure containing the native, redox active site conclusively establish the existence of the Cys-42 sulfenic acid moiety as the functional nonflavin redox center of the peroxidase and provide the first structure for any naturally-occurring protein-sulfenic acid.
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Stehle, T., Ahmed, S.A., Claiborne, A., and Schultz, G.E. (1991). J. Mol. Biol. 221, 1325- 1344.