An ABC Transporter of Streptococcus pneumoniae Involved in Susceptibility to Vancoresmycin and Bacitracin

Abstract

Vancoresmycin is a novel tetramic acid antibiotic, probably interfering with functions of the cytoplasmic membrane. To investigate its mode of action, mutants of Streptococcus pneumoniae exhibiting reduced susceptibility to vancoresmycin were isolated at a low frequency. Four of them were further examined and showed similar pleiotropic phenotypes, including reduced growth rate, early autolysis, and chain formation. In one mutant, the level of transcripts from a single locus encoding the potential ABC transporter Spr0812-Spr0813 was increased sixfold. The corresponding DNA sequence revealed a nonsense mutation (C1744T) in spr0813, leading to the formation of a truncated permease lacking 2 of the 10 predicted transmembrane helices. As demonstrated by deletion and transformation analysis and reconstructing the spr0813 C1744T mutation in the wild-type background, this nucleotide exchange was sufficient to cause reduced susceptibility to vancoresmycin and higher amounts of spr0812-spr0813 mRNA. Mapping and reporter assays of the cognate promoter P abc showed that spr0812 and spr0813 are cotranscribed with a preceding small gene and that the spr0813 C1744T mutation does not affect the activity of P abc . Due to the similarity of Spr0812-Spr0813 to bacitracin transporters of Streptococcus mutans and Bacillus spp., the bacitracin susceptibility of spr0813 mutants was examined. Both the spr0813 C1744T nonsense mutation and the deletion of the transporter genes led to a clearly increased sensitivity to bacitracin. Thus, the intact transporter is required for intrinsic resistance to bacitracin, whereas reduced vancoresmycin susceptibility is mediated by the truncated permease.Antibiotic-resistant bacterial pathogens are of growing concern worldwide. Thus, extensive and ongoing efforts are required to screen for new drugs active against clinically relevant bacteria such as Streptococcus pneumoniae, one of the major human pathogens. Derivatives of tetramic acid (2,4-pyrrolidinedione) represent one group of chemical compounds with a high potential of antibacterial activity (28); however, there is currently no drug of this type in the market. A number of tetramic acid derivatives with antibacterial activity have been detected by testing natural or or chemically synthesized products (see reference 37 [and references therein] and references 24 and 31).The antimicrobial mechanism of action for most of these compounds is not well characterized. Notable exceptions are reutericyclin (produced by Lactobacillus reuteri), which acts as a proton ionophore, dissipating the transmembrane proton potential (4, 5), and several submicromolar inhibitors of undecaprenyl pyrophosphate synthase that were recently designed on the basis of a pharmacophore hypothesis (24). This points to the cytoplasmic membrane as the cellular target of tetramic acid antibiotics and is consistent with the observation that their activity can be improved by introducing more lipophilic side chains at the N-substituted position (37). Partitioning of such compounds into...

Coevolution of ABC Transporters and Two-Component Regulatory Systems as Resistance Modules against Antimicrobial Peptides in Firmicutes Bacteria

“…This is the same TCS identified previously, albeit without experimental evidence, as the most likely regulator of spr0812-spr0813 expression (Fig. 6B) (5). Thus, the phylogenetic classification presented here provides a useful tool for the prediction of likely candidate systems for investigations into the regulatory mechanisms of such orphan transporters.…”

“…Additionally, producer self-resistance against lantibiotics is often mediated by ATP-binding cassette (ABC) transporters, collectively termed LanFEG, consisting of two membrane-spanning subunits and one ATPase, which are encoded in the biosynthetic loci for the respective lantibiotic and whose expression is regulated by a two-component system (TCS) of the same genetic locus (21). Over the last decade, several ABC transporters of a different type have been identified as resistance determinants against peptide antibiotics in nonproducing strains (5,12,32,35,39,41,44,51). The permeases of these transporters share unique domain architecture with 10 transmembrane helices and a large extracellular domain (ECD) of about 200 amino acids between helices 7 and 8.…”

The ABC Transporter AnrAB Contributes to the Innate Resistance of Listeria monocytogenes to Nisin, Bacitracin, and Various β-Lactam Antibiotics

“…Interestingly, a somewhat related ABC transporter in S. pneumoniae D39 (Sp0912-Sp0913; 52% and 26% identity to AnrA and -B, respectively) was recently found to contribute to the innate resistance of this strain to nisin, bacitracin, gramicidin (all are cell envelopeactive antimicrobials), and lincomycin (protein synthesis inhibitor). The other distant homologues of AnrB which have previously been linked to antimicrobial resistance include BceB, MbrB (both linked to bacitracin resistance [6,44]), YsaB, YvsC (both linked to nisin resistance [21,26]), and SPR0813 (linked to resistance to bacitracin and the novel tetrameric antibiotic vancoresmycin [4]). Notably, all of these homologues are members of the ABC M_9 subfamily (www-abcdb.biotoul .fr), whose members are thought to all be exporters due to the lack of associated substrate binding proteins (13).…”

Generic and Specific Adaptive Responses ofStreptococcus pneumoniaeto Challenge with Three Distinct Antimicrobial Peptides, Bacitracin, LL-37, and Nisin

“…The finding that SP0912-0913 is involved in resistance to antimicrobial compounds acting on the cell envelope, i.e., nisin and bacitracin, and antimicrobial compounds involved in protein synthesis inhibition, i.e., lincomycin (10), strongly suggests that the ABC transporter might be of the MDR type. Recently, Becker et al showed that this ABC transporter is indeed involved in resistance of S. pneumoniae R6 to bacitracin (3).…”