Allosteric Enhancement of Adaptational Demethylation by a Carboxyl-terminal Sequence on Chemoreceptors

• Published in: The Journal of biological chemistry 2002-11, Vol.277 (44), p.42151-42156
• Main Authors: Barnakov, Alexander N, Barnakova, Ludmila A, Hazelbauer, Gerald L
• Format: Article
• Language: English
• Subjects: Allosteric Regulation; Amino Acid Sequence; Bacterial Proteins - metabolism; Catalysis; Chemoreceptor Cells - chemistry; Chemoreceptor Cells - metabolism; Dealkylation; Escherichia coli Proteins - metabolism; Kinetics; Membrane Proteins - metabolism; Methyl-Accepting Chemotaxis Proteins; Receptors, Cell Surface - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M206245200

Sensory adaptation in bacterial chemotaxis is mediated by covalent modification of chemoreceptors. Specific glutamyl residues are methylated and demethylated in reactions catalyzed by methyltransferase CheR and methylesterase CheB. In Escherichia coli and Salmonella enterica serovar typhimurium, efficient adaptational modification by either enzyme is dependent on a conserved pentapeptide sequence at the chemoreceptor carboxyl terminus, a position distant from the sites of modification. For CheR-catalyzed methylation, previous work demonstrated that this sequence acts as a high affinity docking site, enhancing methylation by increasing enzyme concentration near methyl-accepting glutamates. We investigated pentapeptide-mediated enhancement of CheB-catalyzed demethylation and found it occurred by a distinctly different mechanism. Assays of binding between CheB and the pentapeptide sequence showed that it was too weak to have a significant effect on local enzyme concentration. Kinetic analyses revealed that interaction of the sequence and the methylesterase enhanced the rate constant of demethylation not the Michaelis constant. This allosteric activation occurred if the sequence was attached to chemoreceptor, but hardly at all if it was present as an isolated peptide. In addition, free peptide inhibited demethylation of the native receptor carrying the pentapeptide sequence at its carboxyl terminus. These observations imply that the allosteric change is transmitted through the protein substrate, not the enzyme.