Cloning and characterization of an exoinulinase from Bacillus polymyxa.
A gene encoding an exoinulinase (inu) from Bacillus polymyxa MGL21 was cloned and sequenced. It is composed of 1455 nucleotides, encoding a protein (485 amino acids) with a molecular mass of 55,522 Da. Inu was expressed in Escherichia coli and the His-tagged exoinulinase was purified. The purified enzyme hydrolyzed sucrose, levan and raffinose, in addition to inulin, with a sucrose/inulin ratio of 2. Inulinase activity was optimal at 35 degrees C and pH 7, was completely inactivated by 1 mM Ag+ or Hg2+. The Km and Vmax values for inulin hydrolysis were 0.7 mM and 2500 microM min(-1) mg(-1) protein. The enzyme acted on inulin via an exo-attack to produce fructose mainly.
Kwon
Hyun-Ju
HJ
Mitsubishi Kagaku Institute of Life Sciences, Tokyo 194-8511, Japan.
Jeon
Sung-Jong
SJ
You
Dong-Ju
DJ
Kim
Kwang-Hyeon
KH
Jeong
Yong-Kee
YK
Kim
Young-Hee
YH
Kim
Young-Man
YM
Kim
Byung-Woo
BW
D000595:Amino Acid Sequence; D001407:Bacillus / Q000145:classification / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification*; D003001:Cloning, Molecular; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005632:Fructose / Q000096:biosynthesis; D006026:Glycoside Hydrolases / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D007444:Inulin / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D016415:Sequence Alignment; D012988:Soil Microbiology; D013045:Species Specificity; D013379:Substrate Specificity; D043324:beta-Fructofuranosidase
D011994:Recombinant Proteins; D005632:Fructose; D007444:Inulin; D006026:Glycoside Hydrolases; D043324:beta-Fructofuranosidase