Cloning and characterization of two genes from Bacillus polymyxa expressing beta-glucosidase activity in Escherichia coli.
DNA fragments from Bacillus polymyxa which encode beta-glucosidase activity were cloned in Escherichia coli by selection of yellow transformants able to hydrolyze the artificial chromogenic substrate p-nitrophenyl-beta-D-glucopyranoside. Restriction endonuclease maps and Southern analysis of the cloned fragments showed the existence of two different genes. Expression of either one of these genes allowed growth of E. coli in minimal medium with cellobiose as the only carbon source. One of the two enzymes was found in the periplasm of E. coli, hydrolyzed arylglucosides more actively than cellobiose, and rendered glucose as the only product upon cellobiose hydrolysis. The other enzyme was located in the cytoplasm, was more active toward cellobiose, and hydrolyzed this disaccharide, yielding glucose and another, unidentified compound, probably a phosphorylated sugar.
González-Candelas
L
L
Instituto de Agroquímica y Tecnología de Alimentos, Consejo Superior de Investigaciones Científicas, Valencia, Spain.
Aristoy
M C
MC
Polaina
J
J
Flors
A
A
D001407:Bacillus / Q000201:enzymology / Q000235:genetics*; D015139:Blotting, Southern; D002475:Cellobiose / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D015964:Gene Expression Regulation, Bacterial; D005959:Glucosidases / Q000235:genetics*; D006868:Hydrolysis; D010957:Plasmids; D015183:Restriction Mapping; D013379:Substrate Specificity; D001617:beta-Glucosidase / Q000235:genetics* / Q000378:metabolism
D004269:DNA, Bacterial; D002475:Cellobiose; D005959:Glucosidases; D001617:beta-Glucosidase