Cloning and immunolocalization of an antifungal chitinase in jelly fig (Ficus awkeotsang) achenes.

ABSTRACT

A 30-kDa protein extracted from the pericarpial portion of jelly fig (Ficus awkeotsang Makino) achenes has been identified as a thermostable chitinase based on its enzymatic activity. A cDNA fragment encoding the precursor protein (including a cleavable signal sequence) of this chitinase was obtained by PCR cloning, and subsequently confirmed by immunological recognition of its overexpressed protein in Escherichia coli. Homology modeling predicted that this thermostable chitinase in jelly fig achenes comprised a stable (betaalpha)(8) barrel fold with three pairs of disulfide linkage. Immunostaining indicated that this chitinase was exclusively localized in the pericarpial region but not in the seed cells where bulky protein bodies and massive oil bodies were accumulated. Spore germination of Colletotrichum gloeosporioides, a common post-harvest pathogen infecting ripening fruit of jelly fig and many other fruits, was inhibited by this chitinase purified from achenes. It is suggested that the biological function of the thermostable chitinase in the pericarp of jelly fig achenes is to protect the nutritive seeds from fungal attack during fruit ripening.