Purification and characterization of alpha-glucosidase I from Japanese honeybee (Apis cerana japonica) and molecular cloning of its cDNA.
Biosci Biotechnol Biochem, 2006/12;70(12):2889-98.
Wongchawalit J[1], Yamamoto T, Nakai H, Kim YM, Sato N, Nishimoto M, Okuyama M, Mori H, Saji O, Chanchao C, Wongsiri S, Surarit R, Svasti J, Chiba S, Kimura A
Affiliations
PMID: 17151473
Impact factor: 2.337
Abstract
alpha-Glucosidase (JHGase I) was purified from a Japanese subspecies of eastern honeybee (Apis cerana japonica) as an electrophoretically homogeneous protein. Enzyme activity of the crude extract was mainly separated into two fractions (component I and II) by salting-out chromatography. JHGase I was isolated from component I by further purification procedure using CM-Toyopearl 650M and Sephacryl S-100. JHGase I was a monomeric glycoprotein (containing 15% carbohydrate), of which the molecular weight was 82,000. Enzyme displayed the highest activity at pH 5.0, and was stable up to 40 degrees C and in a pH-range of 4.5-10.5. JHGase I showed unusual kinetic features: the negative cooperative behavior on the intrinsic reaction on cleavage of sucrose, maltose, and p-nitrophenyl alpha-glucoside, and the positive cooperative behavior on turanose. We isolated cDNA (1,930 bp) of JHGase I, of which the deduced amino-acid sequence (577 residues) confirmed that JHGase I was a member of alpha-amylase family enzymes. Western honeybees (Apis mellifera) had three alpha-glucosidase isoenzymes (WHGase I, II, and III), in which JHGase I was considered to correspond to WHGase I.
MeSH terms
Animals; Base Sequence; Bees; Carbohydrate Metabolism; Chromatography, Gel; Cloning, Molecular; DNA Primers; DNA, Complementary; Electrophoresis, Polyacrylamide Gel; Enzyme Stability; Hydrogen-Ion Concentration; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Substrate Specificity; Temperature; alpha-Glucosidases
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