PMID:1547277
| Citation |
Krueger, JK, Stock, J and Schutt, CE (1992) Evidence that the methylesterase of bacterial chemotaxis may be a serine hydrolase. Biochim. Biophys. Acta 1119:322-6 |
|---|---|
| Abstract |
CheB, the methylesterase of chemotactic bacteria, catalyzes the hydrolysis of glutamyl-methyl esters in bacterial chemoreceptor proteins. The two cysteines predicted by the amino acid sequence of CheB were replaced by alanine residues. The resulting mutants, Cys207-Ala, Cys309-Ala and a double cysteine mutant Cys207-Ala/Cys309-Ala, retained methylesterase activity, indicating that sulfhydryls are not crucial for CheB mediated catalysis. A homology search revealed a conserved serine active-site region between residues 162 and 166 which is homologous to the active-site region of acetylcholine esterases, suggesting that Ser164 of CheB is the active-site nucleophile. Oligonucleotide-directed mutagenesis was used to change the serine to a cysteine. This Ser164-Cys mutant had less than 2% of the wild-type activity. Unlike the serine proteinases which utilize a 'catalytic triad' mechanism, CheB does not have the conserved histidine and aspartic acid residues located in positions N-terminal to the active-site serine. In addition, CheB is not labeled with di-isopropylfluorophosphate, a potent inhibitor of other serine hydrolases. A novel mechanism is proposed for CheB involving substrate-assisted catalysis to account for these apparent anomalies. |
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| Keywords |
Amino Acid Sequence; Bacterial Proteins/chemistry; Bacterial Proteins/genetics; Bacterial Proteins/metabolism; Base Sequence; Binding Sites; Carboxylic Ester Hydrolases/chemistry; Carboxylic Ester Hydrolases/genetics; Carboxylic Ester Hydrolases/metabolism; Chemotactic Factors/chemistry; Chemotactic Factors/genetics; Chemotactic Factors/metabolism; Chemotaxis; Escherichia coli/genetics; Isoflurophate/metabolism; Molecular Sequence Data; Serine/metabolism; Serine Endopeptidases/metabolism |
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