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S-selective hydroxynitrile lyase from a plant Baliospermum montanum: Molecular characterization of recombinant enzyme
Dadashipour, Mohammad1,2; Yamazaki, Mizue1,2; Momonoi, Kazumi3; Tamura, Ken'ichirou1,2; Fuhshuku, Ken-ichi1,2; Kanase, Yurina1,2; Uchimura, Etsuzoh4; Guan Kaiyun5; Asano, Yasuhisa1,2
Corresponding AuthorAsano, Y (reprint author), Toyama Prefectural Univ, Biotechnol Res Ctr, 5180 Kurokawa, Toyama 9390398, Japan
2011-05-20
Source PublicationJOURNAL OF BIOTECHNOLOGY
ISSN0168-1656
Volume153Issue:3-4Pages:100-110
AbstractA novel S-hydroxynitrile lyase (HNL) was purified from leaves of a plant, Baliospermum montanum, by ammonium sulfate fractionation and column chromatographies. Full-length cDNA and genomic DNA were cloned and sequenced. The latter contained two introns and one ORF encoding a 263-residue protein (subunit: 29.5 kDa). The hnl gene was expressed in Escherichia coli and the enzyme was characterized including detailed kinetic studies of 20 substrates for (S)-cyanohydrin synthesis. The enzyme exhibited the highest specific activity (178 U/mg), k(cat) (98/s) and k(cat)/K(m) ratio for piperonal. kcat/Km ratio for aromatic aldehydes was much larger than those of aliphatic aldehydes and ketones. It was strongly inhibited by AgNO(3), PMSF, phenol and methyl ethyl ketone, showed an optimum at pH 5, while having activity at range of 4-6.5. It exhibited stability at wide pH range 2.4-11, the highest activity at 20 degrees C, being active at 0-65 degrees C. The enzyme showed variations in residues involved in substrate pocket and substrate entrance channel compared to other S-selective HNLs, based on a model was built. C-terminal short truncations provided more enzyme production. Gel filtration revealed a 60-65 kDa molecular mass for this non-FAD enzyme and its C-terminally truncated forms using three buffer compositions, indicating dimeric structures. (C) 2011 Elsevier B. V. All rights reserved.
KeywordS-selective Hydroxynitrile Lyase Substrate Specificity Characterization Molecular Cloning Homology Modeling Alanine Scanning
Subject AreaBiotechnology & Applied Microbiology
DOI10.1016/j.jbiotec.2011.02.004
Indexed BySCI
Language英语
WOS Research AreaBiotechnology & Applied Microbiology
WOS SubjectBiotechnology & Applied Microbiology
WOS IDWOS:000290513700004
Citation statistics
Cited Times:24[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.kib.ac.cn/handle/151853/5235
Collection资源植物与生物技术所级重点实验室
Affiliation1.Toyama Prefectural Univ, Biotechnol Res Ctr, Toyama 9390398, Japan
2.Toyama Prefectural Univ, Dept Biotechnol, Toyama 9390398, Japan
3.Toyama Agr Res Ctr, Toyama 9398153, Japan
4.Bot Gardens Toyama, Toyama 9392713, Japan
5.Chinese Acad Sci, Kunming Inst Bot, Kunming Bot Garden, Kunming 650204, Yunnan, Peoples R China
Recommended Citation
GB/T 7714
Dadashipour, Mohammad,Yamazaki, Mizue,Momonoi, Kazumi,et al. S-selective hydroxynitrile lyase from a plant Baliospermum montanum: Molecular characterization of recombinant enzyme[J]. JOURNAL OF BIOTECHNOLOGY,2011,153(3-4):100-110.
APA Dadashipour, Mohammad.,Yamazaki, Mizue.,Momonoi, Kazumi.,Tamura, Ken'ichirou.,Fuhshuku, Ken-ichi.,...&Asano, Yasuhisa.(2011).S-selective hydroxynitrile lyase from a plant Baliospermum montanum: Molecular characterization of recombinant enzyme.JOURNAL OF BIOTECHNOLOGY,153(3-4),100-110.
MLA Dadashipour, Mohammad,et al."S-selective hydroxynitrile lyase from a plant Baliospermum montanum: Molecular characterization of recombinant enzyme".JOURNAL OF BIOTECHNOLOGY 153.3-4(2011):100-110.
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