Cloning and Amplified Expression in Streptomyces Iividans of the Gene Encoding the Extracellular β-Lactamase from Streptumyces cacaoi

SUMMARY: A 19 kb SphI DNA fragment containing the gene for the extracellular active-site serine β-lactamase of Streptomyces cacaoi KCC-SO352 was cloned in Streptomyces liuidans TK24 using the high-copy-number plasmid pIJ702 as vector. A 30-fold higher yield of β-lactamase was obtained from S. lividans strain ML1, carrying the recombinant plasmid pDML51, than from S. cacaoi grown under optimal production conditions. In all respects (molecular mass, isoelectric point, kinetics of inhibition by β-iodopenicillanate) the overproduced S. lividans ML 1 β-lactamase was identical to the original S. cacaoi enzyme. A considerable reduction of β-lactamase production was caused by elimination of a 12.8 kb portion of the 19 kb DNA fragment by cleavage at an internal SphI site located more than 3 kb upstream of the β-lactamase structural gene. The β-lactamase gene was located within a 1.8 NcoI-BclI fragment but when this fragment was cloned in S. lividans pIJ702, the resulting strain produced hardly any more β-lactamase than the original S. cacaoi.