Molecular cloning and characterization of two lincomycin‐resistance genes, ImrA and ImrB, from Streptomyces lincolnensls 78–11

Abstract

Two different lincomycin-resistance determinants (lmrA and lmrB) from Streptomyces lincolnensis 78-11 were cloned in Streptomyces lividans 66 TK23. The gene lmrA was localized on a 2.16 kb fragment, the determined nucleotide sequence of which encoded a single open reading frame 1446 bp long. Analysis of the deduced amino acid sequence suggested the presence of 12 membrane-spanning domains and showed significant similarities to the methylenomycin-resistance protein (Mmr) from Streptomyces coelicolor, the QacA protein from Staphylococcus aureus, and several tetracycline-resistance proteins from both Gram-positive and Gram-negative bacteria, as well as to some sugar-transport proteins from Escherichia coli. The lmrB gene was actively expressed from a 2.7 kb fragment. An open reading frame of 837 bp could be localized which encoded a protein that was significantly similar to 23S rRNA adenine(2058)-N-methyltransferases conferring macrolide-lincosamide-streptogramin resistance. LmrB also had putative rRNA methyltransferase activity since lincomycin resistance of ribosomes was induced in lmrB-containing strains. Surprisingly, both enzymes, LmrA and LmrB, had a substrate specificity restricted to lincomycin and did not cause resistance to other lincosamides such as celesticetin and clindamycin, or to macrolides.

The pur8 gene from the pur cluster of Streptomyces alboniger encodes a highly hydrophobic polypeptide which confers resistance to puromycin

“…Transmembrane proteins involved in transport and/or drug resistance contain multiple (12)(13)(14) transmembrane regions [6,41]. These similarities are extended to four partially conserved motifs which are present in Pur8 at positions 49 and 50, 82-94, 117-129 and 163-172 [6,331. The @-turn motif [RKI-X-G-R- [RK] (X, unknown amino acid), which is found between transmembrane domains 2 and 3 in most eukaryotic and prokaryotic transport proteins [41,421, is localized at positions 87-92 of Pur8 (Fig.…”

“…The relationships and possible evolutionary origin of the transport proteins with similarities to Pur8 have been considered at length elsewhere [6,41,431. Briefly, these transmembrane polypeptides are thought to have evolved from a common ancestor and share a similar amino-terminal region, which appears to be directly implicated in general transport functions that, as shown for tetracycline-resistance proteins [44], would be energized by a proton-dependent electrochemical gradient .…”

“…However, as has been noted for several of those proteins by Marger and Saier [41], Pur8 does not exhibit full conservation of this motif, since the first residue is alanine. Similarly, this position is changed to leucine in LmrA [6].…”

“…The highest levels of similarity, extending over the complete sequence, existed with presumptive transmembrane proteins from antibiotic-producing Streptornyces, including LmrA from S. lincolnensis [6], TcmA from S. glaucescens [4] and Mmr from S. coelicolor [S], which confer resistance to lincomycin, tetracenomycin C and methylenomycin, respectively (Figs 4 and 5). In addition, a significant level of similarity was found with QacA, which confers resistance to a variety of antiseptic and desinfectant agents in Staphylococcus uureus [33] and for two proteins from S. coelicolor, ActII-ORF2 and ActV-ORF1, which appear to be implicated in the efflux of actinorhodin [2, …”

“…posed by Zhang et al [6] for LmrA, Mmr and QacA in addition to other groups of transmembrane transporter proteins [41 I.…”