Regulation of 2-deoxyglucose phosphate accumulation in Lactococcus lactis vesicles by metabolite-activated, ATP-dependent phosphorylation of serine-46 in HPr of the phosphotransferase system.
Lactococcus lactis takes up glucose and the nonmetabolizable glucose analogue 2-deoxyglucose (2DG) via the phosphotransferase system and extrudes the accumulated sugar phosphates in a process apparently dependent on a cytoplasmic sugar-phosphate phosphatase. Uptake of 2DG into L. lactis vesicles was shown to be dependent on an energy source, effectively provided by intravesicular phosphoenolpyruvate (PEP). 2DG phosphate (2DG-P) accumulation in these vesicles was not inhibited, and preaccumulated 2DG-P was not released from them, upon electroporation of fructose 1,6-diphosphate (FDP), gluconate 6-phosphate or 2-phosphoglycerate into the vesicles. Intravesicular but not extravesicular wild-type HPr of Bacillus subtilis alone stimulated uptake, but in the presence of any one of these metabolites, it prevented accumulation of 2DG-P. Intravesicular H15A mutant HPr inhibited uptake and allowed further inhibition of 2DG-P accumulation in the presence of the intravesicular metabolites. Intravesicular S46A mutant HPr stimulated uptake but could not promote inhibition in the presence of the phosphorylated metabolites. The S46D mutant HPr protein promoted regulation, even in the absence of a metabolite. The Vmax but not the Km value for 2DG uptake was affected. Accumulation of the natural, metabolizable substrates of the lactose, glucose, mannose and ribose permeases was inhibited by wild-type HPr in the presence of FDP or by S46D mutant HPr. The results establish that HPr serine phosphorylation by the ATP-dependent, metabolite-activated HPr kinase selectively determines the levels of sugar accumulation via the glucose and lactose permeases in L. lactis.(ABSTRACT TRUNCATED AT 250 WORDS)
Ye
J J
JJ
Department of Biology, University of California at San Diego, La Jolla 92093-0116.
Reizer
J
J
Saier
M H
MH
D000255:Adenosine Triphosphate / Q000378:metabolism; D001693:Biological Transport, Active; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000493:pharmacokinetics; D003847:Deoxyglucose / Q000378:metabolism; D005958:Glucosephosphates / Q000378:metabolism*; D007700:Kinetics; D013294:Lactococcus lactis / Q000235:genetics / Q000378:metabolism*; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000235:genetics; D017354:Point Mutation; D012694:Serine / Q000378:metabolism
D002241:Carbohydrates; D005958:Glucosephosphates; D012694:Serine; C081196:2-deoxyglucose-1-phosphate; D000255:Adenosine Triphosphate; D003847:Deoxyglucose; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System