Characterization of the KdpD protein, the sensor kinase of the K+-translocating Kdp system of Escherichia coli

Abstract

KdpD and KdpE, proteins that control expression of the kdpFABC operon, are members of the class of sensor kinase/response regulator proteins. Using polyclonal antibodies raised against the KdpD protein, we have been able to identify and to localize the chromosome-encoded KdpD protein in the cytoplasmic membrane of Escherichia coli. Furthermore, it has been possible to detect differences in the expression of the KdpD protein according to the K+ concentration in the growth medium. The phosphorylation capacity of the plasmid-encoded KdpD protein and the phospho-transfer to KdpE was investigated. We found that both reactions were strictly dependent on the ionic conditions of the assay medium. Based on optimized conditions, we were able to detect phosphorylation of the chromosome-encoded KdpD protein. Furthermore, replacement of the conserved histidine (His673), the predicted phosphorylation site in KdpD, by glutamine revealed that phosphorylation of KdpD was no longer possible.