OLIGOMERIZATION OF THE PHOSPHATASE CHEZ UPON INTERACTION WITH THE PHOSPHORYLATED FORM OF CHEY - THE SIGNAL PROTEIN OF BACTERIAL CHEMOTAXIS

Earlier studies have suggested that CheZ, the phosphatase of the signa ling protein CheY in bacterial chemotaxis, may be in an oligomeric sta te both when bound to phosphorylated CheY (CheY similar to P) (Blat, Y ., and Eisenbach, M. (1994) Biochemistry 33, 902-906) or free (Stock, A., and Stock, J. B. (1987) J. Bacteriol. 169, 3301-3311). The purpose of the current study was to determine the oligomeric state of free Ch eZ and to investigate whether it changes upon binding to CheY similar to P. By using either one of two different sets of cross-linking agent s, free CheZ was found to be a dimer. The formation of the dimer was s pecific, as it was prevented by SDS which does not interfere with cros s-linking mediated by random collisions. The dimeric form of CheZ was confirmed by sedimentation analysis, a cross-linking-free technique. I n the presence of CheY similar to P (but not in the presence of non-ph osphorylated CheY), a high molecular size crosslinked complex (90-200 kDa) was formed, in which the CheZ:CheY ratio was 2:1. The size of the oligomeric complex was estimated by fluorescence depolarization to be 4-5-fold larger than the dimer, suggesting that its size is in the or der of 200 kDa. These results indicate that CheZ oligomerizes upon int eraction with CheY similar to P3. This phosphorylation-dependent oligo merization may be a mechanism for regulating CheZ activity.