Cloning, expression, and characterization of an antifungal chitinase from Leucaena leucocephala de wit

被引：30

作者：

Kaomek, M

Mizuno, K

Fujimura, T

Sriyotha, P

Cairns, JRK ^{[1
]}

机构：

[1] Suranaree Univ Technol, Inst Sci, Nakhon Ratchasima 30000, Thailand

[2] Rajabhat Inst Pecthburiwittayalongkorn, Fac Sci & Technol, Pathum Thani 13180, Thailand

[3] Univ Tsukuba, Inst Agr & Forest Engn, Tsukuba, Ibaraki 3058572, Japan

[4] Chulabhorn Res Inst, Biochem Lab, Bangkok 10210, Thailand

来源：

BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY | 2003年 / 67卷 / 04期

关键词：

chitinases; Leucaena leucocephala de Wit; cloning; recombinant expression;

D O I：

10.1271/bbb.67.667

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

Chitinase cDNAs from Leucaena leucocephala seedlings were cloned by PCR amplification with degenerate primers based on conserved class I chitinase sequences and cDNA library screening. Two closely related chitinase cDNAs were sequenced and inferred to encode precursor proteins of 323 (KB1) and 326 (KB2) amino acids. Expression of the KB2 chitinase from a pET32a plasmid in Origami (DE3) Escherichia coli produced high chitinase activity in the cell lysate. The recombinant thioredoxin fusion protein was purified and cleaved to yield a 32-kDa chitinase. The recombinant chitinase hydrolyzed colloidal chitin with endochitinase-type activity. It also inhibited growth of 13 of the 14 fungal strains tested.

引用

页码：667 / 676

页数：10

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