Molecular cloning and characterization of a class III chitinase in pumpkin leaves, which strongly binds to regenerated chitin affinity gel

A class' III chitinase (PL-ChtIII) with a molecular weight of 29 kDa was purified to homogeneity from pumpkin leaves using regenerated chitin affinity gel and high-performance liquid chromatography Mono-Q anion exchange column chromatography. In contrast to other class III chitinases reported in plants, the PL-ChtIII,strongly binds to regenerated chitin gel column, which can provide a convenient tool for the enzyme purification. An oligonucleotide probe derived from the N-terminal amino acid sequence of the purified PL-ChtIII was used in cloning the cDNA. The full-length cDNA of the PL-ChtIII clone consists of 27 and 128 binding pairs (bp) of 5'- and 3'-untranslated region, respectively, and 864 bp of open reading frame. The deduced amino acid sequence of the clone shares significant homology with plant class III chitinase enzymes. The nucleotide sequence of the PL-ChtIII cDNA predicts the synthesis of a preprotein, which is subsequently processed into a mature protein by removal of an N-terminal signal peptide. The protein contains six cysteine residues conserved in all class III chitinases at their invariant positions. Expression of PL-ChtIII mRNA is significantly induced within 1-3 h by the treatment of fungal elicitor or glycol chitin, but the expression of PL-ChtIII protein maximally occurred 12 h after the elicitor treatments. (C) 1999 Elsevier Science Ireland Ltd. All rights reserved.
Publisher
ELSEVIER SCI IRELAND LTD
Issue Date
1999-09
Language
ENG
Keywords

CYSTEINE-RICH DOMAIN; PLANT CHITINASES; BEAN-LEAVES; WHEAT-GERM; ETHYLENE; PROTEINS; GENES; ENDOCHITINASE

Citation

PLANT SCIENCE, v.147, no.2, pp.157 - 163

ISSN
0168-9452
URI
http://hdl.handle.net/10203/74802
Appears in Collection
BS-Journal Papers(저널논문)
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