Gene Cloning and Characterization of a Novel Recombinant Antifungal Chitinase from Papaya (Carica papaya)

A chitinase cDNA clone (CpCHI, 1002 bp) was isolated from papaya fruit, which encoded a 275 amino acid protein containing a 28 amino acid signal peptide in the N-terminal end. The predicted molecular mass of the mature protein was 26.2 kDa, and its pI value was 6.32. On the basis of its amino acid s...

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Bibliographic Details
Published in:Journal of agricultural and food chemistry 2007-02, Vol.55 (3), p.714-722
Main Authors: Chen, Yu-Ting, Hsu, Lien-Hua, Huang, I-Ping, Tsai, Tsun-Chuang, Lee, Guan-Chiun, Shaw, Jei-Fu
Format: Article
Language:English
Subjects:
Alternaria - drug effects
Alternaria brassicicola
Amino Acid Sequence
antibacterial properties
antifungal properties
Base Sequence
Biological and medical sciences
Carica - enzymology
Carica papaya
chitinase
Chitinases - chemistry
Chitinases - genetics
Chitinases - pharmacology
complementary DNA
enzyme activity
Escherichia coli
Escherichia coli - genetics
Food industries
Fruit - enzymology
Fruit and vegetable industries
Fundamental and applied biological sciences. Psychology
Fungicides, Industrial - pharmacology
Hydrogen-Ion Concentration
isoelectric point
molecular cloning
Molecular Sequence Data
molecular weight
papayas
recombinant proteins
Recombinant Proteins - chemistry
Recombinant Proteins - genetics
Recombinant Proteins - pharmacology
spore germination
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Summary:A chitinase cDNA clone (CpCHI, 1002 bp) was isolated from papaya fruit, which encoded a 275 amino acid protein containing a 28 amino acid signal peptide in the N-terminal end. The predicted molecular mass of the mature protein was 26.2 kDa, and its pI value was 6.32. On the basis of its amino acid sequence homology with other plant chitinases, it was classified as a class IV chitinase. An active recombinant CpCHI enzyme was overexpressed in Escherichia coli. The purified recombinant papaya chitinase showed an optimal reaction temperature at 30 °C and a broad optimal pH ranging from 5.0 to 9.0. The recombinant enzyme was quite stable, retaining >64% activity for 3 weeks at 30 °C. The spore germination of Alternaria brassicicola could be completely inhibited by a 76 nM level of recombinant CpCHI. Recombinant CpCHI also showed antibacterial activity in which 50% of E. coli was inhibited by a 2.5 μM concentration of the enzyme. Keywords: Chitinase; papaya; antifungal activity; recombinant expression
ISSN:0021-8561
1520-5118
DOI:10.1021/jf062453e