RRC ID |
28618
|
著者 |
Nakajima M, Yamashita T, Takahashi M, Nakano Y, Takeda T.
|
タイトル |
Identification, cloning, and characterization of β-glucosidase from Ustilago esculenta.
|
ジャーナル |
Appl Microbiol Biotechnol
|
Abstract |
Hydrolytic enzymes responsible for laminarin degradation were found to be secreted during growth of Ustilago esculenta on laminarin. An enzyme involved in laminarin degradation was purified by assaying release of glucose from laminaribiose. Ion-exchange chromatography of the culture filtrate followed by size-exclusion chromatography yielded a 110-kDa protein associated with laminaribiose hydrolysis. LC/MS/MS analysis of the 110-kDa protein identified three peptide sequences that shared significant similarity with a putative glucoside hydrolase family (GH) 3 β-glucosidase in Ustilago maydis. Based on the DNA sequence of the U. maydis GH3 β-glucosidase, a gene encoding a putative GH3 β-glucosidase in U. esculenta (Uebgl3A) was cloned by PCR. Based on the deduced amino acid sequence, the protein encoded by Uebgl3A has a molecular mass of 91 kDa and shares 90% identity with U. maydis GH3 β-glucosidase. Recombinant UeBgl3A expressed in Aspergillus oryzae released glucose from β-1,3-, β-1,4-, and β-1,6-linked oligosaccharides, and from 1,3-1,4-β-glucan and laminarin polysaccharides, indicating that UeBgl3A is a β-glucosidase. Kinetic analysis showed that UeBgl3A preferentially hydrolyzed laminaritriose and laminaritetraose. These results suggest that UeBgl3A is a key enzyme that produces glucose from laminarioligosaccharides during growth of U. esculenta on laminarin.
|
巻・号 |
93(5)
|
ページ |
1989-98
|
公開日 |
2012-3-1
|
DOI |
10.1007/s00253-011-3538-2
|
PMID |
21850431
|
MeSH |
Aspergillus oryzae / enzymology
Aspergillus oryzae / genetics
Chromatography, Gel
Chromatography, Ion Exchange
Cloning, Molecular
Glucans
Molecular Weight
Phylogeny
Polysaccharides / metabolism
Recombinant Proteins / chemistry
Recombinant Proteins / genetics
Recombinant Proteins / metabolism
Sequence Homology, Amino Acid
Tandem Mass Spectrometry
Ustilago / enzymology*
Ustilago / genetics
beta-Glucosidase / chemistry
beta-Glucosidase / genetics
beta-Glucosidase / metabolism*
|
IF |
3.53
|
引用数 |
18
|
WOS 分野
|
BIOTECHNOLOGY & APPLIED MICROBIOLOGY
|
リソース情報 |
遺伝子材料 |
pCR8/GW/TOPO-UeBgl3A (RDB08966) |