title: Characterization of trehalose phosphorylase from Bacillus stearothermophilus SK-1 and nucleotide sequence of the corresponding gene.
abstract: A bacterial trehalose phosphorylase (TPase; EC 2.4.1.64) was purified from the culture supernatant of Bacillus stearothermophilus SK-1 to apparent homogeneity, and some properties were investigated. Furthermore, a gene from SK-1 responsible for the TPase was cloned by Southern hybridization with a degenerate oligonucleotide probe synthesized on the basis of the N-terminal sequence of the purified enzyme. The Mr of the enzyme was estimated to be 150,000 by gel filtration and 83,000 by SDS-PAGE, so the enzyme is likely to be a homodimer. The enzyme had optimum activity at pH 7.0-8.0 or nearby and the optimum temperature was about 75 degrees C. The deduced amino acid sequence of the SK-1 TPase encodes a theoretical protein with a Mr of 87,950. Alignment of amino acid sequences with a maltose phosphorylase from Lactobacillus brevis the crystal structure and active site of which had been analyzed suggested that these two phosphorylases evolved from a common ancestor. The Escherichia coli cells harboring the plasmid containing the cloned TPase gene had about 100 times the activity of SK-1.
authors.affiliation: Showa Sangyo Co, Ltd, Tsukubashi, Ibaraki, Japan. brc@showa-sangyo.co.jp
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Inoue
authors.forename: Keiko
authors.initials: K
authors.lastname: Ishii
authors.forename: Tetsuji
authors.initials: T
authors.lastname: Tomita
authors.forename: Tsuneya
authors.initials: T
authors.lastname: Yatake
authors.forename: Fumio
authors.initials: F
authors.lastname: Fukui
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D004926:Escherichia coli; D001411:Geobacillus stearothermophilus / Q000201:enzymology* / Q000235:genetics; D005964:Glucosyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D005964:Glucosyltransferases; C023181:alpha,alpha-trehalose phosphorylase