title: The fructose transporter of Bacillus subtilis encoded by the lev operon: backbone assignment and secondary structure of the IIB(Lev) subunit.
abstract: The fructose transporter of the Bacillus subtilis phosphotransferase system consists of two membrane associated (IIA and IIB) and two transmembrane (IIC and IID) subunits [Martin-Verstraete, I., Débarbouille, M., Klier, A. & Rapoport, G. (1990) J. Mol. Biol. 214, 657-671] . It mediates uptake by a mechanism which couples translocation to phosphorylation of the transported solute. The 18-kDa IIBLev subunit transfers phosphoryl groups from His9 of the IIA subunit to the sugar. The three-dimensional structure of IIBLev or similar proteins is not known. IIBLev was overexpressed in Escherichia coli and isotopically labelled with 13C/15N in H2O as well as in 70% D2O. 15N-edited NOESY, 13C-edited NOESY and 13C,15N triple-resonance experiments yielded a nearly complete assignment of the 1H, 13C and 15N resonances. Based on qualitative interpretation of NOE, scalar couplings, chemical shift values and amide exchange data, the secondary structure and topology of IIBLev was determined. IIBLev comprises six parallel beta-strands, one antiparallel beta-strand and 5 alpha-helices. The order of the major secondary-structure elements is (beta alpha)5beta (strand order 7651423). Assuming that the (beta alpha beta)-motives form right-handed turn structures, helices alphaA and alphaB are packed to one face and helices alphaC, alphaD and alphaE to the opposite face of the parallel beta-sheet. His15 which is transiently phosphorylated during catalysis is located in the loop beta1/alphaA of the topological switch point. The amino terminal (beta/alpha)4 part of IIBLev has the same topology as phosphoglyceromutase (PGM; PDB entry 3pgm). Both proteins catalyze phosphoryltransfer reactions which proceed through phosphohistidine intermediates and they show a similar distribution of invariant residues in the topologically equivalent positions of their active sites. The protein fold of IIBLev has no similarity to any of the known structures of other phosphoenolpyruvate-dependent-carbohydrate-phosphotransferase-system proteins.
authors.affiliation: PH-R-Structural Research, Bayer AG, Wuppertal, Germany. debayrj9@ibmmail.com
authors.forename: S
authors.initials: S
authors.lastname: Seip
authors.forename: R
authors.initials: R
authors.lastname: Lanz
authors.forename: R
authors.initials: R
authors.lastname: Gutknecht
authors.forename: K
authors.initials: K
authors.lastname: Flükiger
authors.forename: B
authors.initials: B
authors.lastname: Erni
mesh_terms: D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics; D001693:Biological Transport, Active; D006860:Hydrogen Bonding; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000737:chemistry; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000737:chemistry* / Q000235:genetics; D009876:Operon; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000737:chemistry* / Q000235:genetics*; D010736:Phosphoglycerate Mutase / Q000737:chemistry; D010766:Phosphorylation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins
chemical_list: D008565:Membrane Proteins; D009004:Monosaccharide Transport Proteins; D011994:Recombinant Proteins; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C022409:protein-N-pi-phosphohistidine - sugar phosphotransferase; D010736:Phosphoglycerate Mutase