Abstract
A clone library of hybrid plasmids containing DNA from the chitinolytic marine bacterium Vibrio harveyi was constructed in Escherichia coli. Three of 2,450 clones capable of expressing chitobiase activity were selected by their ability to hydrolyze p-nitrophenyl-2-acetamido-2-deoxy-β-D-glucopyranoside. Additional testing confirmed the expression of chitobiase activity by two of these E. coli clones, which were further analyzed for the production of chitinase. Both of these clones, carrying inserts 5.3 kb and 13.5 kb in length, exhibited chitinase activity as well, indicating that the chitinase and chitobiase genes are linked in V. harveyi. Induction studies with V. harveyi revealed that N,N'-diacetylchitobiose is a strong inducer of both enzyme activities. In the E. coli clones, however, both genes are constantly expressed, and the activities of both enzymes remain unchanged regardless of whether or not N,N'-diacetylchitobiose is added to the growth medium. Our evidence suggests that there is a chi operon in V. harveyi consisting of genes coding for chitinase, chitobiase, and possibly a permease for N,N'-diacetylchitobiose, and that the entire operon is present in two of our clones.
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