Linked Life Data

11139129

Source:http://linkedlifedata.com/resource/pubmed/id/11139129

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
3-4
pubmed:dateCreated
2001-1-2
pubmed:abstractText
Random mutagenesis has been developed as an approach for the study of human cytochrome P450 (P450) enzymes and their structure and function. Sensitive screening methods are critical for the success of this approach. We have developed one system that takes advantage of the ability of human P450 1A2 to activate heterocyclic amines to mutagenic products [A. Parikh, P. D. Josephy, and F. P. Guengerich, Biochemistry, 38, 5283-5289 (1999)]. Mutants with both attenuated and enhanced activity have been recovered and subjected to further kinetic analysis. For phenacetin O-deethylation, the E225I mutant had kcat 6x > wild type; D320A had kcat 1/10x < wild type (and Km 15 x > wild type). With all three P450s, the rate of first electron reduction was similar, and all had similar binding constants for phenacetin (approximately 15 microM). All three forms yielded intermolecular, noncompetitive kinetic deuterium isotope effects of 1.5-2 [DV and D(V/K)] for O-deethylation of [OCD2CH3]-phenacetin. All three forms of P450 1A2 also formed a minor product, the acetol (C-hydroxylation of the acetyl group). This reaction had a deuterium isotope effect of approximately 14 with all three forms of the enzyme, and C-H bond breaking is the rate-determining step. Another approach to P450 2A6 involves the recent observation that this P450 can accumulate indigo [E. M. J. Gillam, A. M. A. Aguinaldo, L. M. Notley, D. Kim, R. G. Mundkowski, A. A. Volkov, F. H. Arnold, P. Soucek, J. T. DeVoss, and F. P. Guengerich, Biochem. Biophys. Res. Commun 265, 469-472 (1999)]. Current results indicate that this process involves the conversion of endogenous indole to indoxyl by the P450. The reaction may be used in assays of random mutants and has some potential applications in industry.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:issn
0360-2532
pubmed:author
pubmed:issnType
Print
pubmed:volume
32
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
267-81
pubmed:dateRevised
2011-11-17
pubmed:meshHeading
pubmed:articleTitle
What makes P450s work? Searches for answers with known and new P450s.
pubmed:affiliation
Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0146, USA. guengerich@toxicology.mc.vanderbilt.edu
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Review, Research Support, Non-U.S. Gov't