Open AccessCloning and sequencing of a gene encoding the 69‐kDa extracellular chitinase of Janthinobacterium lividum
Author(s) -
Gleave Andrew P.,
Taylor Robert K.,
Morris Bret A.M.,
Greenwood David R.
Publication year - 1995
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1995.tb07788.x
Subject(s) - chitinase , biology , biochemistry , peptide sequence , microbiology and biotechnology , open reading frame , nucleic acid sequence , amino acid , gene
Janthinobacterium lividum secretes a major 56‐kDa chitinase and a minor 69‐kDa chitinase. A chitinase gene was defined on a 3‐kb fragment of clone pRKT10, by virtue of fluorescent colonies in the presence of 4‐methylumbelliferyl‐β‐d‐N,N′,N″‐chitotrioside. Nucleotide sequencing revealed an 1998‐bp open reading frame with the potential to encode a 69 716‐Da protein with amino acid sequences similar to those in other chitinases, suggesting it encodes the minor chitinase (Chi69). Chitinase activity of Escherichia coli (pRKTIO) lysates was detected mainly in the periplasmic fraction and immunoblotting detected a 70‐kDa protein in this fraction. Chi69 has an N‐terminal secretory leader peptide preceding two probable chitin‐binding domains and a catalytic domain. These functional domains are separated by linker regions of proline‐threonine repeats. Amino acid sequencing of cyanogen bromide cleavage‐derived peptides from the major 56‐kDa chitinase suggested that Chi69 may be a precursor of Chi56. In addition, an N‐terminally truncated version of Chi69 retained chitinase activity as expected if in vivo processing of Chi69 generates Chi56.