Molecular characterization and transcriptional analysis of a multidrug resistance gene cloned from the pristinamycin-producing organism, Streptomyces pristinaespiralis.
A multidrug resistance gene (mdr) has been cloned from Streptomyces pristinaespiralis, a producer of two antibiotics having synergistic activities together known as pristinamycin. This gene, ptr, provides resistance not only to two structurally dissimilar compounds (pristinamycin I, PI; pristinamycin II, PII) and the natural pristinamycin mixture but also to rifampicin. Mutagenesis and subcloning of ptr localized it to a 2 kb region which was sequenced and analyzed. It contained an open reading frame of 1506 bp which encoded a putative membrane protein with 14 hydrophobic domains, and showed sequence similarity to a superfamily of bacterial proteins that employ transmembrane electrochemical gradients to catalyse active efflux of various antibiotics and toxic compounds. Ptr was most similar to a subfamily which included other mdr genes and antibiotic transport genes associated with antibiotic biosynthetic gene clusters in actinomycetes. In vitro coupled transcription-translation experiments were used to identify the ptr gene product. Analysis of the upstream region did not reveal a divergently transcribed repressor gene, as is the case for several related resistance determinants involved in antibiotic transport, suggesting that ptr is regulated by a different mechanism. Transcriptional analyses of this gene, carried out in both S. pristinaespiralis and Streptomyces lividans, indicated the same transcriptional start point and predicted -10 and -35 hexamers which were somewhat similar to Streptomyces vegetative-type promoters.
Blanc
V
V
Institut Pasteur, Paris, France.
Salah-Bey
K
K
Folcher
M
M
Thompson
C J
CJ
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000096:biosynthesis* / Q000494:pharmacology; D001483:Base Sequence; D003001:Cloning, Molecular; D018432:Drug Resistance, Multiple / Q000235:genetics*; D005798:Genes, Bacterial; D008565:Membrane Proteins / Q000096:biosynthesis; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016366:Open Reading Frames; D010957:Plasmids; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis; D015183:Restriction Mapping; D013302:Streptomyces / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D014769:Virginiamycin / Q000096:biosynthesis*
D000900:Anti-Bacterial Agents; D008565:Membrane Proteins; D012329:RNA, Bacterial; D011994:Recombinant Proteins; D014769:Virginiamycin