Journal: mBio
Article Title: An Aspartate-Specific Solute-Binding Protein Regulates Protein Kinase G Activity To Control Glutamate Metabolism in Mycobacteria
doi: 10.1128/mBio.00931-18
Figure Lengend Snippet: PknG activation by glutamate was indirect, and PknG function required the conserved protein interaction domain (TPR domain). (A) Kinase activity of recombinant PknG toward GarA was independent of glutamate. Kinase reactions in the presence or absence of glutamate were sampled at the indicated times. Phosphorylation of GarA was detected by the known decrease in mobility of phospho-GarA in SDS-PAGE ( , ). The rate of GarA phosphorylation was similar to that seen in previous studies of PknG and was unchanged by the addition of glutamate. Images are representative of three independent experiments. Similar results were obtained with other putative activators: glutamine, aspartate, and α-ketoglutarate. (B) PknG homologues from Actinomycetales have a variable N-terminal domain, sometimes lack the Rdx domain (absent from homologues in the Corynebacteriaceae ), and always have a tetratricopeptide repeat (TPR) domain. (C) Truncated PknG lacking the Rdx was able to restore the ability of pknG -deficient M. smegmatis to grow on minimal medium containing glutamate as the sole nitrogen source, but truncated PknG lacking the TPR domain did not improve growth. Error bars show standard deviations from three wells, and the growth curve is representative of three independent experiments.
Article Snippet: TPR domains have been identified in kinases from multiple bacterial phyla (>1,000 listed in Interpro [ ]), suggesting that TPR-mediated protein interaction could be a widespread regulatory mechanism of bacterial serine/threonine protein kinases.
Techniques: Activation Assay, Activity Assay, Recombinant, Phospho-proteomics, SDS Page
