In vitro phosphorylation of key metabolic enzymes from Bacillus subtilis: PrkC phosphorylates enzymes from different branches of basic metabolism.
Phosphorylation is an important mechanism of protein modification. In the Gram-positive soil bacterium Bacillus subtilis, about 5% of all proteins are subject to phosphorylation, and a significant portion of these proteins is phosphorylated on serine or threonine residues. We were interested in the regulation of the basic metabolism in B. subtilis. Many enzymes of the central metabolic pathways are phosphorylated in this organism. In an attempt to identify the responsible protein kinase(s), we identified four candidate kinases, among them the previously studied kinase PrkC. We observed that PrkC is indeed able to phosphorylate several metabolic enzymes in vitro. Determination of the phosphorylation sites revealed a remarkable preference of PrkC for threonine residues. Moreover, PrkC often used several phosphorylation sites in one protein. This feature of PrkC-dependent protein phosphorylation resembles the multiple phosphorylations often observed in eukaryotic proteins. The HPr protein of the phosphotransferase system is one of the proteins phosphorylated by PrkC, and PrkC phosphorylates a site (Ser-12) that has recently been found to be phosphorylated in vivo. The agreement between in vivo and in vitro phosphorylation of HPr on Ser-12 suggests that our in vitro observations reflect the events that take place in the cell.
Pietack
Nico
N
Department of General Microbiology, Institute of Microbiology and Genetics, Georg August University, Göttingen, Germany.
Becher
Dörte
D
Schmidl
Sebastian R
SR
Saier
Milton H
MH
Hecker
Michael
M
Commichau
Fabian M
FM
Stülke
Jörg
J
D001412:Bacillus subtilis / Q000201:enzymology*; D001426:Bacterial Proteins / Q000302:isolation & purification* / Q000378:metabolism*; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000302:isolation & purification / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000302:isolation & purification / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D012694:Serine / Q000378:metabolism; D013912:Threonine / Q000378:metabolism
D001426:Bacterial Proteins; D013912:Threonine; D012694:Serine; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C030182:phosphocarrier protein HPr; D011493:Protein Kinase C