Journal
CHEMICAL RESEARCH IN TOXICOLOGY
Volume 18, Issue 8, Pages 1203-1210Publisher
AMER CHEMICAL SOC
DOI: 10.1021/tx050100o
Keywords
-
Funding
- NCI NIH HHS [CA16954] Funding Source: Medline
Ask authors/readers for more resources
Tyr190 in cytochrome P450 2131 has previously been shown to be a prime target for nitration by peroxynitrite (PN) resulting in nitrotyrosine formation and the inactivation of this enzyme. Modeling studies have suggested that Tyr190 may play a structural role in maintaining the integrity of the protein for maximal activity through hydrogen bonding with Glu149. To elucidate the roles of Tyr190 and Glu149 hydrogen-bonding in maintaining the catalytically competent structure of P450 2131, we have mutated Tyr190 to Phe or Ala and Glu149 to Gln or Ala to characterize the catalytic activities and the structural stabilities of mutated proteins. The results demonstrate that (a) the catalytic activities of all four mutants were decreased significantly compared to wild-type (WT); (b) nitration of Tyr190 by PN or mutation of Tyr190 to Phe did not alter the Km of the reductase for P450; (c) PN decreases the catalytic activity of the heat-treated Y190A, E149Q, and E149A mutants to a much greater extent than the WT and Y190F; and (d) after exposure of the P450s to PN, the extent of nitrotyrosine formation and the inactivation of the catalytic activity of the E149Q and E149A mutants were markedly decreased when compared to WT. These findings suggest that (1) the hydrogen bond between Tyr190 and Glu149 stabilizes the protein for maximal activity; (2) the benzyl ring and hydroxyl groups of Tyr190 stabilize the protein structure when P450 is exposed to the temperatures higher than 45 degrees C; and (3) Glu149 may be critical in directing the site of nitration by PN. Since Glu149 and Tyr190 are both highly conserved in the P450 2 family, they may play an important role in the tertiary structure and functional properties of these P450s.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available