title: Purification and partial biochemical-genetic characterization of trehalose 6-phosphate synthase from muscles of adult female Ascaris suum.
abstract: Trehalose 6-phosphate (T6P) synthase (TPS; EC 2.4.1.15) was isolated from muscles of Ascaris suum by ammonium sulphate fractionation, ion-exchange DEAE SEPHACEL(TM) anion exchanger column chromatography and Sepharose 6B gel filtration. On sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), 265-fold purified TPS exhibited a molecular weight of 66 kDa. The optimum pH and temperature of the purified enzyme were 3.8-4.2 and 35°C, respectively. The isoelectric point (pI) of TPS was pH 5.4. The studied TPS was not absolutely substrate specific. Besides glucose 6-phosphate, the enzyme was able to use fructose 6-phosphate as an acceptor of glucose. TPS was activated by 10 mM MgCl2, 10 mM CaCl2 and 10 mM NaCl. In addition, it was inhibited by ethylenediaminetetra-acetic acid (EDTA), KCl, FeCl3 and ZnCl2. Two genes encoding TPS were isolated and sequenced from muscles of the parasite. Complete coding sequences for tps1 (JF412033.2) and tps2 (JF412034.2) were 3917 bp and 3976 bp, respectively. Translation products (AEX60788.1 and AEX60787.1) showed expression to the glucosyltransferase-GTB-type superfamily.
authors.affiliation: Department of Biochemistry, Faculty of Biology, University of Warmia and Mazury, Olsztyn, Poland. m.dmit@uwm.edu.pl
authors.forename: M
authors.initials: M
authors.lastname: Dmitryjuk
authors.forename: M
authors.initials: M
authors.lastname: Dopieralska
authors.forename: E
authors.initials: E
authors.lastname: Łopieńska-Biernat
authors.forename: R J
authors.initials: RJ
authors.lastname: Frączek
mesh_terms: D000818:Animals; D017165:Ascaris suum / Q000201:enzymology*; D005591:Chemical Fractionation; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D018509:DNA, Helminth / Q000737:chemistry / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D020536:Enzyme Activators / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D004795:Enzyme Stability; D005260:Female; D005964:Glucosyltransferases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology; D010802:Phylogeny; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D018509:DNA, Helminth; D020536:Enzyme Activators; D004791:Enzyme Inhibitors; D005964:Glucosyltransferases; C023180:trehalose-6-phosphate synthase