Regulatory interactions of a virulence-associated serine/threonine phosphatase-kinase pair in Bacillus anthracis.
In the current study, we examined the regulatory interactions of a serine/threonine phosphatase (BA-Stp1), serine/threonine kinase (BA-Stk1) pair in Bacillus anthracis. B. anthracis STPK101, a null mutant lacking BA-Stp1 and BA-Stk1, was impaired in its ability to survive within macrophages, and this correlated with an observed reduction in virulence in a mouse model of pulmonary anthrax. Biochemical analyses confirmed that BA-Stp1 is a PP2C phosphatase and dephosphorylates phosphoserine and phosphothreonine residues. Treatment of BA-Stk1 with BA-Stp1 altered BA-Stk1 kinase activity, indicating that the enzymatic function of BA-Stk1 can be influenced by BA-Stp1 dephosphorylation. Using a combination of mass spectrometry and mutagenesis approaches, three phosphorylated residues, T165, S173, and S214, in BA-Stk1 were identified as putative regulatory targets of BA-Stp1. Further analysis found that T165 and S173 were necessary for optimal substrate phosphorylation, while S214 was necessary for complete ATP hydrolysis, autophosphorylation, and substrate phosphorylation. These findings provide insight into a previously undescribed Stp/Stk pair in B. anthracis.
Shakir
Salika M
SM
University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.
Bryant
Katie M
KM
Larabee
Jason L
JL
Hamm
Elaine E
EE
Lovchik
Julie
J
Lyons
C Rick
CR
Ballard
Jimmy D
JD
D000818:Animals; D001408:Bacillus anthracis / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D015153:Blotting, Western; D002460:Cell Line; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D013058:Mass Spectrometry; D051379:Mice; D008807:Mice, Inbred BALB C; D016297:Mutagenesis, Site-Directed; D004676:Myelin Basic Protein / Q000378:metabolism; D010748:Phosphopeptides / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014774:Virulence / Q000235:genetics / Q000502:physiology*
D001426:Bacterial Proteins; D004676:Myelin Basic Protein; D010748:Phosphopeptides; D017346:Protein Serine-Threonine Kinases; D010749:Phosphoprotein Phosphatases