Molecular analysis of the phosphoenolpyruvate-dependent L-sorbose: phosphotransferase system from Klebsiella pneumoniae and of its multidomain structure.
We have cloned a 3.4 kb DNA fragment from the chromosome of Klebsiella pneumoniae that codes for a phosphoenolpyruvate-dependent L-sorbose: phosphotransferase system (PTS). The cloned fragment was sequenced and four open reading frames coding for 135 (sorF), 164 (sorB), 266 (sorA) and 274 (sorM) amino acids, respectively, were found. The corresponding proteins could be detected in a T7 overexpression system, which yielded molecular masses of about 14,000 for SorF, 19,000 for SorB, 25,000 for SorA and 27,000 for SorM. SorF and SorB have all the characteristics of soluble and intracellular proteins in accordance with their functions as EIIASor and EIIBSor domains of the L-sorbose PTS. SorA and SorM, by contrast, are strongly hydrophobic, membrane-bound proteins with two to five putative transmembrane helices that alternate with a series of hydrophilic loops. They correspond to domains EIICSor and EIIDSor. The four proteins of the L-sorbose PTS resemble closely (27%-60%) the four subunits of a D-fructose PTS (EIIALev, EIIBLev, EIICLev, and EIIDLev) from Bacillus subtilis and the three subunits of the D-mannose PTS (EIIA,BMan, EIICMan, and EIIDMan) from Escherichia coli K-12. The three systems constitute a new PTS family, and sequence comparisons revealed highly conserved structures for the membrane-bound proteins. A consensus sequence for the membrane proteins was used to postulate a model for their integration into the membrane.
Wehmeier
U F
UF
Bergische Universität-Gesamthochschule Wuppertal, Chemische Mikrobiologie, Germany.
Wöhrl
B M
BM
Lengeler
J W
JW
D000595:Amino Acid Sequence; D001483:Base Sequence; D001693:Biological Transport, Active; D002462:Cell Membrane / Q000378:metabolism; D004790:Enzyme Induction; D005632:Fructose / Q000494:pharmacology; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D007711:Klebsiella pneumoniae / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D009876:Operon / Q000235:genetics; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000737:chemistry / Q000235:genetics*; D011487:Protein Conformation; D015183:Restriction Mapping; D013013:Sorbose / Q000378:metabolism* / Q000494:pharmacology
D005632:Fructose; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C067377:phosphoenolpyruvate-sorbose phosphotransferase; D013013:Sorbose