Cloning and analysis of the beta-lactamase gene from epsilon-poly-L-lysine-producing actinomycete Streptomyces albulus IFO14147.
Streptomyces albulus IFO14147 produces epsilon-poly-L-lysine, which exhibits antimicrobial activity. It is necessary for its molecular breeding to develop host-vector systems. We recently found a novel cryptic plasmid, pNO33, in this strain. As part of a search for a selectable marker gene for pNO33, we report here the isolation and analysis of the beta-lactamase gene of this strain, which can grow on ampicillin-containing plates. It was shown that the beta-lactamase production in S. albulus was induced by ampicillin. By introducing a genomic library of S. albulus into Escherichia coli, a 3.6-kbp fragment was identified as the region involved in ampicillin resistance. It contained three open reading frames, all of which are highly homologous to the beta-lactamase (the blaL product) and its regulatory proteins (the blaA and blaB products) of S. cacaoi. The growth phenotypes and enzyme assaying of E. coli and S. lividans showed that the blaL homologue (blaSa) encodes a beta-lactamase required for ampicillin resistance. The beta-lactamae gene can be utilized as a selectable marker in a cloning vector of S. albulus. However, the beta-lactamase activity was decreased in E. coli and repressed in S. lividans by the blaA and blaB homologues (blaASa and blaBSa). It appears as if the blaASa product is a repressor of blaSa instead of an activator as in S. cacaoi.
Hoshino
Yusuke
Y
Department of Bioscience, Fukui Prefectural University, 4-1-1 Kenjojima, Matsuoka-cho, Fukui 910-1195.
Nakamori
Shigeru
S
Takagi
Hiroshi
H
D000667:Ampicillin / Q000494:pharmacology; D000668:Ampicillin Resistance; D003001:Cloning, Molecular; D003470:Culture Media / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D010957:Plasmids; D013302:Streptomyces / Q000235:genetics* / Q000378:metabolism; D013997:Time Factors; D001618:beta-Lactamases / Q000235:genetics*
D003470:Culture Media; D000667:Ampicillin; D001618:beta-Lactamases