Characterization of an HPr kinase mutant of Staphylococcus xylosus.
The Staphylococcus xylosus gene hprK, encoding HPr kinase (HPrK), has been isolated from a genomic library. The HPrK enzyme, purified as a His(6) fusion protein, phosphorylated HPr, the phosphocarrier protein of the bacterial phosphotransferase system, at a serine residue in an ATP-dependent manner, and it also catalyzed the reverse reaction. Therefore, the enzyme constitutes a bifunctional HPr kinase/phosphatase. Insertional inactivation of the gene in the genome of S. xylosus resulted in the concomitant loss of both HPr kinase and His serine-phosphorylated-HPr phosphatase activities in cell extracts, strongly indicating that the HPrK enzyme is also responsible for both reactions in vivo. HPrK deficiency had a profound pleiotropic effect on the physiology of S. xylosus. The hprK mutant strain showed a severe growth defect in complex medium upon addition of glucose. Glucose uptake in glucose-grown cells was strongly enhanced compared with the wild type. Carbon catabolite repression of three tested enzyme activities by glucose, sucrose, and fructose was abolished. These results clearly demonstrate the prominent role of HPr kinase in global control to adjust catabolic capacities of S. xylosus according to the availability of preferred carbon sources.
Huynh
P L
PL
Mikrobielle Genetik, Universität Tübingen, D-72076 Tübingen, Germany.
Jankovic
I
I
Schnell
N F
NF
Brückner
R
R
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001483:Base Sequence; D050260:Carbohydrate Metabolism; D003001:Cloning, Molecular; D015964:Gene Expression Regulation, Bacterial; D005947:Glucose / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000378:metabolism; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000235:genetics; D009154:Mutation / Q000235:genetics*; D010641:Phenotype; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D011765:Pyruvaldehyde / Q000378:metabolism; D012329:RNA, Bacterial / Q000032:analysis / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013210:Staphylococcus / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics
D001426:Bacterial Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D011765:Pyruvaldehyde; D000255:Adenosine Triphosphate; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C030182:phosphocarrier protein HPr; C110847:HPr kinase; D017346:Protein Serine-Threonine Kinases; C110846:YvoE protein, Bacillus substilis; D010749:Phosphoprotein Phosphatases; D006026:Glycoside Hydrolases; D005947:Glucose