Document Detail

Heme oxygenase active-site residues identified by heme-protein cross-linking during reduction of CBrCl3.
MedLine Citation:
PMID:  9485440     Owner:  NLM     Status:  MEDLINE    
The reduction of CBrCl3 by the heme-heme oxygenase complex forms dissociable and covalently bound heme products. No such products are formed with mesoheme in which the heme vinyl substituents are replaced by ethyl groups. The dissociable heme products are chromatographically similar but not identical to those obtained in the analogous reaction with myoglobin. Tryptic digestion of the heme-protein adduct and Edman sequencing and mass spectrometric analysis of the heme-linked peptide identify His-25, the proximal iron ligand, as the alkylated residue. Reaction of CBrCl3 with the heme complexes of the T135V mutant and a Delta221 C-terminal truncated protein yields heme-linked peptides in addition to that from the wild-type reaction. The sequence of the principal labeled peptide from the T135V reaction, 205TAFLLNIQLFEELQELLTHDTK226 , and the lability of the adduct suggest the heme is attached to one of the carboxylic acid residues. A carboxylic acid residue is probably also labeled in the modified peptide 49LVMASLYHIYVALEEEIER67 from the Delta221 truncated protein. Thus, addition of the reductively generated trichloromethyl radical to a heme vinyl group produces a species that alkylates active-site residues. The changes in the alkylated residue caused by the Thr-135 mutation or truncation of the protein places residues in the sequences 49-67 and 205-226 within the active site. Furthermore, this is the first demonstration that heme oxygenase, like cytochrome P450, may catalyze the reductive metabolism of halocarbons and thus contribute to the toxicity of these agents.
A Wilks; K F Medzihradszky; P R Ortiz de Montellano
Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Biochemistry     Volume:  37     ISSN:  0006-2960     ISO Abbreviation:  Biochemistry     Publication Date:  1998 Mar 
Date Detail:
Created Date:  1998-04-03     Completed Date:  1998-04-03     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  0370623     Medline TA:  Biochemistry     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  2889-96     Citation Subset:  IM    
Department of Pharmaceutical Chemistry, School of Pharmacy, and Liver Center, University of California, San Francisco, California 94143-0446, USA.
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MeSH Terms
Amino Acid Sequence
Binding Sites
Bromotrichloromethane / chemistry,  metabolism*
Chromatography, High Pressure Liquid
Cross-Linking Reagents
Heme / chemistry,  metabolism*
Heme Oxygenase (Decyclizing) / chemistry,  metabolism*
Molecular Sequence Data
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Structure-Activity Relationship
Grant Support
Reg. No./Substance:
0/Cross-Linking Reagents; 14875-96-8/Heme; 75-62-7/Bromotrichloromethane; EC Oxygenase (Decyclizing)

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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