Regulation of the activity of the Bacillus subtilis antiterminator LicT by multiple PEP-dependent, enzyme I- and HPr-catalysed phosphorylation.
The transcriptional antiterminator LicT regulates the induction and carbon catabolite repression of the Bacillus subtilis bglPH operon. LicT is inactive in mutants affected in one of the two general components of the phosphoenolpyruvate (PEP):glycose phosphotransferase system, enzyme I or histidine-containing protein (HPr). We demonstrate that LicT becomes phosphorylated in the presence of PEP, enzyme I and HPr. The phosphoryl group transfer between HPr and LicT is reversible. Phosphorylation of LicT with PEP, enzyme I and HPr led to the appearance of three additional LicT bands on polyacrylamide-urea gels. These bands probably correspond to one-, two- and threefold phosphorylated LicT. After phosphorylation of LicT with [32P]-PEP, enzyme I and HPr, proteolytic digestion of [32P]-P-LicT, separation of the peptides by reverse-phase chromatography, mass spectrometry and N-terminal sequencing of radiolabelled peptides, three histidyl residues were found to be phosphorylated in LicT. These three histidyl residues (His-159, His-207 and His-269) are conserved in most members of the BglG/SacY family of transcriptional antiterminators. Phosphorylation of LicT in the presence of serylphosphorylated HPr (P-Ser-HPr) was much slower compared with its phosphorylation in the presence of HPr. The slower phosphorylation in the presence of P-Ser-HPr leading to reduced LicT activity is presumed to play a role in a recently described LicT-mediated CcpA-independent carbon catabolite repression mechanism operative for the bglPH operon.
Institut de Biologie et Chimie des Protéines, Centre National de la Recherche Scientifique, UPR412, Lyon, France.
C
C
Lindner
A
A
Galinier
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M
Hecker
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J
Deutscher
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000235:genetics* / Q000502:physiology; D001426:Bacterial Proteins / Q000235:genetics*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000032:analysis / Q000502:physiology; D010766:Phosphorylation; D017852:Phosphotransferases (Nitrogenous Group Acceptor) / Q000502:physiology; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics*
D001426:Bacterial Proteins; C098955:LicT protein, Bacteria; D014157:Transcription Factors; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C031629:phosphoenolpyruvate-glucose phosphotransferase; D017852:Phosphotransferases (Nitrogenous Group Acceptor); C022360:phosphoenolpyruvate-protein phosphotransferase