The first α-1,3-glucosidase from bacterial origin belonging to glycoside hydrolase family 31

• Published in: Biochimie Vol. 91; no. 11; pp. 1434 - 1442
• Main Authors: Kang, Min-Sun, Okuyama, Masayuki, Mori, Haruhide, Kimura, Atsuo
• Format: Journal Article
• Language: English
• Published: France Elsevier Masson SAS 01.11.2009
• Subjects: alpha-Glucosidases; Cloning, Molecular; Disaccharides - chemistry; Disaccharides - metabolism; DNA, Bacterial; Escherichia coli - enzymology; Escherichia coli - genetics; Glycoside hydrolase family 31; Glycoside Hydrolases - metabolism; Inclusion body formation; Lactobacillus johnsonii; Molecular Sequence Data; Potassium Channels, Inwardly Rectifying - genetics; Potassium Channels, Inwardly Rectifying - metabolism; Sequence Alignment; Sequence Homology, Amino Acid; Yeasts - genetics; α-1,3-Glucosidase; UAN; TLC; DP; BSA; Glycoside hydrolase family 31; Lactobacillus johnsonii; rLJAG31; α-1,3-Glucosidase; HPLC; PTLC; PBS; GH; pNPG; Inclusion body formation; ESI-MS; FAB-MS; ER; MalA; SDS-PAGE; pNP; LB; ND; IPTG; TIM; BA
• ISSN: 0300-9084; 1638-6183
• DOI: 10.1016/j.biochi.2009.07.018

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 α-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 α-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 α-1,3-glucosidase to be characterized with a high k cat/ K m value for nigerose [α- d-Glc p-(1 → 3)- d-Glc p]. Transglucosylation of 4-nitrophenyl α- d-glucopyranoside produced two 4-nitrophenyl disaccharides (4-nitrophenyl α-nigeroside and 4-nitrophenyl α-isomaltoside). These hydrolysis and transglucosylation properties of LJAG31 are different from those of mold ( Acremonium implicatum) α-1,3-glucosidase of glycoside hydrolase family 31.