In vitro phosphorylation of AlgR, a regulator of mucoidy in Pseudomonas aeruginosa, by a histidine protein kinase and effects of small phospho‐donor molecules

Abstract

AlgR is a transcriptional regulator of mucoidy in Pseudomonas aeruginosa, a critical virulence factor expressed in cystic fibrosis. AlgR belongs to the superfamily of bacterial signal transduction systems, and has been shown to bind to the algD promoter, a critical point in the regulation of mucoidy. This protein, like other typical response regulators, contains highly conserved residues known to be critical for the phosphorylation and signal transduction processes. However, a typical second component interacting with AlgR has not been identified. Here we demonstrate that AlgR undergoes phosphorylation in vitro when interacting with the well-characterized histidine protein kinase CheA. These results indicate that AlgR is capable of undergoing phosphorylation typical of other two-component signal transduction systems. Moreover, the phosphotransfer reaction between CheA and AlgR was found to be affected by the presence of carbamoyl phosphate, acetyl phosphate, and salts of phosphoramidic acid, recently shown to act as small-molecular-weight phospho-donors in the process of phosphorylation of several response regulators. These findings suggest that AlgR may react with intermediary metabolites such as carbamoyl phosphate and acetyl phosphate, and that these processes may play a role in the control of mucoidy in P. aeruginosa.

The alginate regulator AlgR and an associated sensor FimS are required for twitching motility in Pseudomonas aeruginosa.

“…2b). AlgR has been reported to be capable of being phosphorylated by both histidine kinases, such as CheA, and by small molecular weight phosphate donors, such as carbamoyl phosphate and acetyl phosphate (35). Alternatively, it is possible that FimS blocks or reverses AlgR phosphorylation.…”

Stimulus Perception in Bacterial Signal-Transducing Histidine Kinases

“…The RRs then catalyze their own phosphorylation, with the phosphoryl-HK as the phosphodonor (279,280). In some cases, low-molecular-weight donors such as acetyl phosphate or, in vitro, also phosphoramidates can be used for RR phosphorylation (45,158).…”

Is acetyl phosphate a global signal in Escherichia coli?

“…It has been known for some time that when an autophosphorylated HK protein is incubated with the appropriate unphosphorylated response regulator, transfer of the phosphoryl group to the response regulator occurs (9,40,43 [7,8,16,19]) have been phosphorylated by using one of these three small phosphorylated compounds. Thus, it is apparent that the response regulator proteins catalyze their own phosphorylation, i.e., the phosphotransfer reaction from the HK phosphohistidine to the RRD aspartate target is an autophosphorylation reaction catalyzed by the RRD (35).…”