PMID:9515909
| Citation |
Heath, RJ and Rock, CO (1998) A conserved histidine is essential for glycerolipid acyltransferase catalysis. J. Bacteriol. 180:1425-30 |
|---|---|
| Abstract |
Sequence analysis of membrane-bound glycerolipid acyltransferases revealed that proteins from the bacterial, plant, and animal kingdoms share a highly conserved domain containing invariant histidine and aspartic acid residues separated by four less conserved residues in an HX4D configuration. We investigated the role of the invariant histidine residue in acyltransferase catalysis by site-directed mutagenesis of two representative members of this family, the sn-glycerol-3-phosphate acyltransferase (PlsB) and the bifunctional 2-acyl-glycerophosphoethanolamine acyltransferase/acyl-acyl carrier protein synthetase (Aas) of Escherichia coli. Both the PlsB[H306A] and Aas[H36A] mutants lacked acyltransferase activity. However, the Aas[H36A] mutant retained significant acyl-acyl carrier protein synthetase activity, illustrating that the lack of acyltransferase activity was specifically associated with the H36A substitution. The invariant aspartic acid residue in the HX4D pattern was also important. The substitution of aspartic acid 311 with glutamic acid in PlsB resulted in an enzyme with significantly reduced catalytic activity. Substitution of an alanine at this position eliminated acyltransferase activity; however, the PlsB[D311A] mutant protein did not assemble into the membrane, indicating that aspartic acid 311 is also important for the proper folding and membrane insertion of the acyltransferases. These data are consistent with a mechanism for glycerolipid acyltransferase catalysis where the invariant histidine functions as a general base to deprotonate the hydroxyl moiety of the acyl acceptor. |
| Links | |
| Keywords |
Acyltransferases/genetics; Acyltransferases/metabolism; Alanine/genetics; Amino Acid Sequence; Amino Acid Substitution; Aspartic Acid/genetics; Cell Membrane/metabolism; Cloning, Molecular; DNA, Bacterial/analysis; DNA, Bacterial/genetics; Escherichia coli/genetics; Escherichia coli/metabolism; Gene Expression; Genetic Complementation Test; Glutamic Acid/genetics; Glycerol-3-Phosphate O-Acyltransferase/genetics; Glycerol-3-Phosphate O-Acyltransferase/metabolism; Histidine/genetics; Molecular Sequence Data; Plasmids; Polymerase Chain Reaction; Protein Biosynthesis; Sequence Alignment; Sequence Analysis, DNA; Transcription, Genetic |
| edit table |
Significance
You can help EcoliWiki by summarizing why this paper is useful
Useful Materials and Methods
You can help Ecoliwiki by describing the useful materials (strains, plasmids, antibodies, etc) described in this paper.
Annotations
<annotationlinks/>
EcoliWiki Links
Add links to pages that link here (e.g. gene, product, method pages)
References
See Help:References for how to manage references in EcoliWiki.