The first alpha-1,3-glucosidase from bacterial origin belonging to glycoside hydrolase family 31.
Genome analysis of Lactobacillus johnsonii NCC533 has been recently completed. One of its annotated genes, lj0569, encodes the protein having the conserved domain of glycoside hydrolase family 31. Its homolog gene (ljag31) in L. johnsonii NBRC13952 was cloned and expressed using an Escherichia coli expression system, resulting in poor production of recombinant LJAG31 protein due to inclusion body formation. Production of soluble recombinant LJAG31 was improved with high concentration of NaCl in medium, possible endogenous chaperone induction by benzyl alcohol, and over-expression of GroES-GroEL chaperones. Recombinant LJAG31 was an alpha-glucosidase with broad substrate specificity toward both homogeneous and heterogeneous substrates. This enzyme displayed higher specificity (in terms of k(cat)/K(m)) toward nigerose, maltulose, and kojibiose than other natural substrates having an alpha-glucosidic linkage at the non-reducing end, which suggests that these sugars are candidates for prebiotics contributing to the growth of L. johnsonii. To our knowledge, LJAG31 is the first bacterial alpha-1,3-glucosidase to be characterized with a high k(cat)/K(m) value for nigerose [alpha-d-Glcp-(1 --> 3)-d-Glcp]. Transglucosylation of 4-nitrophenyl alpha-d-glucopyranoside produced two 4-nitrophenyl disaccharides (4-nitrophenyl alpha-nigeroside and 4-nitrophenyl alpha-isomaltoside). These hydrolysis and transglucosylation properties of LJAG31 are different from those of mold (Acremonium implicatum) alpha-1,3-glucosidase of glycoside hydrolase family 31.
Kang
Min-Sun
MS
Division of Applied Bioscience, Research Faculty of Agriculture, Hokkaido University, Kita-9 Nishi-9, Kita-ku, Sapporo 060-8589, Japan. leslie@abs.agr.hokudai.ac.jp
Okuyama
Masayuki
M
Mori
Haruhide
H
Kimura
Atsuo
A
D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004187:Disaccharides / Q000737:chemistry / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D006026:Glycoside Hydrolases / Q000378:metabolism; D008969:Molecular Sequence Data; D024661:Potassium Channels, Inwardly Rectifying / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D015003:Yeasts / Q000235:genetics; D000520:alpha-Glucosidases
D004269:DNA, Bacterial; D004187:Disaccharides; C510692:Kcnj1 protein, rat; D024661:Potassium Channels, Inwardly Rectifying; C014637:laminaribiose; C008237:kojibiose; D006026:Glycoside Hydrolases; D000520:alpha-Glucosidases