title: Isolation and characterization of a novel intracellular glucosyltransferase from the acarbose producer Actinoplanes sp. CKD485-16.
abstract: A novel intracellular glucosyltransferase (GTase) was isolated from cells of Actinoplanes sp. CKD485-16-acarbose-producing cells. The enzyme was purified by DEAE-cellulose and G75-40 Sephadex chromatography. The molecular mass of the enzyme was estimated to be 62 kDa by SDS-polyacrylamide gel electrophoresis, and its isoelectric point (pI) was pH 4.3. The N-terminal sequence of the GTase consisted of NH(2)-Ser-Val-Pro-Leu-Ser-Leu-Pro-Ala-Glu-Trp. The optimum pH and temperature were 7.5 and 30 degrees C. The enzyme was stable in a pH range of 5.5-9.0 and below 40 degrees C. Enzymatic reactions were performed by incubating the GTase with various substrates. The GTase converted acarbose into component C, maltose into trehalose, and maltooligosaccharides into maltooligosyl trehaloses. The reactions were reversible. Various acarbose analogs were tested as inhibitors against the GTase as a means to suppress component C formation. Valienamine was the most potent, with an IC(50) value of 2.4x10(-3) mM and showed a competitive inhibition mode.
authors.affiliation: Department of Biotechnology, College of Engineering, Yonsei University, Seodaemun-gu, Seoul, 120-749, South Korea.
authors.forename: B T
authors.initials: BT
authors.lastname: Choi
authors.forename: C S
authors.initials: CS
authors.lastname: Shin
mesh_terms: D020909:Acarbose / Q000378:metabolism*; D053138:Cyclohexenes; D005964:Glucosyltransferases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D065089:Glycoside Hydrolase Inhibitors; D006595:Hexosamines / Q000494:pharmacology; D007700:Kinetics; D008848:Micromonosporaceae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D013379:Substrate Specificity
chemical_list: D053138:Cyclohexenes; D065089:Glycoside Hydrolase Inhibitors; D006595:Hexosamines; C030791:valienamine; D005964:Glucosyltransferases; D020909:Acarbose