ABC transporter genes, kasKLM, responsible for self-resistance of a kasugamycin producer strain.
We previously reported that a 7.6-kb DNA fragment from Streptomyces kasugaensis M338-M1, a kasugamycin (KSM) producer, included KSM acetyltransferase gene (kac338) and some other genes possibly involved in KSM biosynthesis. As an extension of that study, a 10-kb SacI-KpnI DNA fragment, located approximately 5-15-kb upstream of kac33, was cloned and a 4.2-kb SacI-EcoRI fragment therefrom was sequenced, revealing one incomplete (designated ORF J) and three complete open reading frames (designated kasK, kasL and kasM). The coding frames of kasK, L and M overlap one another with terminator/initiator ATGA sequence. RT-PCR analysis of a DNA region including kasKLM indicated the presence of one transcript that is long enough to span the three genes. The kasK gene potentially encodes an ATP-binding protein of the ATP-binding cassette (ABC) transporter superfamily. Homology search for the deduced KasK protein shows similarity to other ABC transporters involved in self-resistance of a mithramycin and possibly doxorubicin producer strain. The kasL and kasM genes encode different integral membrane proteins, both having six putative transmembrane helices. An expression plasmid for kasKLM (pTV-KLM) was constructed and these genes were expressed in E. coli JM 109, which had been sensitive to KSM. The transformant acquired resistance to KSM, suggesting that KasK, L and M proteins as a set in S. kasugaensis M338-M1 pump out KSM to protect the producer from its toxic metabolite.
Ikeno
S
S
Showa College of Pharmaceutical Sciences, Tokyo, Japan.
Yamane
Y
Y
Ohishi
Y
Y
Kinoshita
N
N
Hamada
M
M
Tsuchiya
K S
KS
Hori
M
M
D018528:ATP-Binding Cassette Transporters / Q000235:genetics*; D000595:Amino Acid Sequence; D000617:Aminoglycosides; D000900:Anti-Bacterial Agents / Q000096:biosynthesis*; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D004269:DNA, Bacterial / Q000235:genetics*; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010957:Plasmids / Q000235:genetics*; D013302:Streptomyces / Q000235:genetics*; D001616:beta-Galactosidase / Q000378:metabolism
D018528:ATP-Binding Cassette Transporters; D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D001616:beta-Galactosidase; C100305:kasugamycin