INTERMOLECULAR COMPLEMENTATION OF THE KINASE-ACTIVITY OF CHEA

CheA is a dimeric autophosphorylating protein kinase that plays a crit ical role in the signal transduction network controlling chemotaxis in Escherichia coli. The autophosphorylation reaction was analysed using mutant proteins defective in kinase and regulatory functions. Protein s in which the site of autophosphorylation was mutated (CheA48HQ) or m issing (CheA(S)) were found to phosphorylate the kinase-defective muta nt, CheA470GK. The kinetics of this reaction support the hypothesis th at autophosphorylation is the result of trans-phosphorylation within a dimer. The carboxy-terminal portion of CheA was previously shown to b e dispensable for autophosphorylation, but required for regulation in response to environmental signals transmitted through a transducer and CheW. Mixing of CheA48HQ or CheA470GK with a truncated protein lackin g this regulatory domain demonstrated that regulated autophosphorylati on requires the presence of both carboxy-terminal portions in a CheA d imer. These results indicate that the dimeric form of CheA plays an in tegral role in signal transduction in bacterial chemotaxis.