Bioscience, Biotechnology, and Biochemistry
Online ISSN : 1347-6947
Print ISSN : 0916-8451
Biochemistry & Molecular Biology Regular Papers
Cloning, Expression, and Characterization of an Antifungal Chitinase from Leucaena leucocephala de Wit
Mana KAOMEKKouichi MIZUNOTatsuhito FUJIMURAPoonsook SRIYOTHAJames R. Ketudat CAIRNS
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2003 Volume 67 Issue 4 Pages 667-676

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Abstract
  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.
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© 2003 by Japan Society for Bioscience, Biotechnology, and Agrochemistry
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