Linked Life Data

15579906

Source:http://linkedlifedata.com/resource/pubmed/id/15579906

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
6
pubmed:dateCreated
2005-2-7
pubmed:abstractText
Carnitine palmitoyltransferase (CPT) I catalyzes the conversion of long-chain fatty acyl-CoAs to acyl carnitines in the presence of l-carnitine, a rate-limiting step in the transport of long-chain fatty acids from the cytoplasm to the mitochondrial matrix. To determine the role of the 15 cysteine residues in the heart/skeletal muscle isoform of CPTI (M-CPTI) on catalytic activity and malonyl-CoA sensitivity, we constructed a 6-residue N-terminal, a 9-residue C-terminal, and a 15-residue cysteineless M-CPTI by cysteine-scanning mutagenesis. Both the 9-residue C-terminal mutant enzyme and the complete 15-residue cysteineless mutant enzyme are inactive but that the 6-residue N-terminal cysteineless mutant enzyme had activity and malonyl-CoA sensitivity similar to those of wild-type M-CPTI. Mutation of each of the 9 C-terminal cysteines to alanine or serine identified a single residue, Cys-305, to be important for catalysis. Substitution of Cys-305 with Ala in the wild-type enzyme inactivated M-CPTI, and a single change of Ala-305 to Cys in the 9-residue C-terminal cysteineless mutant resulted in an 8-residue C-terminal cysteineless mutant enzyme that had activity and malonyl-CoA sensitivity similar to those of the wild type, suggesting that Cys-305 is the residue involved in catalysis. Sequence alignments of CPTI with the acyltransferase family of enzymes in the GenBank led to the identification of a putative catalytic triad in CPTI consisting of residues Cys-305, Asp-454, and His-473. Based on the mutagenesis and substrate labeling studies, we propose a mechanism for the acyltransferase activity of CPTI that uses a catalytic triad composed of Cys-305, His-473, and Asp-454 with Cys-305 serving as a probable nucleophile, thus acting as a site for covalent attachment of the acyl molecule and formation of a stable acyl-enzyme intermediate. This would in turn allow carnitine to act as a second nucleophile and complete the acyl transfer reaction.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Feb
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
11
pubmed:volume
280
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
4524-31
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed-meshheading:15579906-Alanine, pubmed-meshheading:15579906-Amino Acid Sequence, pubmed-meshheading:15579906-Animals, pubmed-meshheading:15579906-Binding Sites, pubmed-meshheading:15579906-Blotting, Western, pubmed-meshheading:15579906-Carnitine, pubmed-meshheading:15579906-Carnitine O-Palmitoyltransferase, pubmed-meshheading:15579906-Catalysis, pubmed-meshheading:15579906-Cysteine, pubmed-meshheading:15579906-DNA Primers, pubmed-meshheading:15579906-Humans, pubmed-meshheading:15579906-Kinetics, pubmed-meshheading:15579906-Malonyl Coenzyme A, pubmed-meshheading:15579906-Models, Chemical, pubmed-meshheading:15579906-Molecular Sequence Data, pubmed-meshheading:15579906-Mutagenesis, pubmed-meshheading:15579906-Mutation, pubmed-meshheading:15579906-Myocardium, pubmed-meshheading:15579906-Palmitic Acid, pubmed-meshheading:15579906-Palmitoylcarnitine, pubmed-meshheading:15579906-Pichia, pubmed-meshheading:15579906-Protein Structure, Tertiary, pubmed-meshheading:15579906-Sequence Homology, Amino Acid, pubmed-meshheading:15579906-Serine
pubmed:year
2005
pubmed:articleTitle
Cysteine-scanning mutagenesis of muscle carnitine palmitoyltransferase I reveals a single cysteine residue (Cys-305) is important for catalysis.
pubmed:affiliation
Department of Environmental and Biomolecular Systems, OGI School of Science and Engineering, Oregon Health & Science University, Beaverton, Oregon 97006-8921, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.