The Pseudomonas aeruginosa PA14 ABC Transporter NppA1A2BCD Is Required for Uptake of Peptidyl Nucleoside Antibiotics

Abstract

Analysis of the genome sequence of Pseudomonas aeruginosa PA14 revealed the presence of an operon encoding an ABC-type transporter (NppA1A2BCD) showing homology to the Yej transporter of Escherichia coli. The Yej transporter is involved in the uptake of the peptide-nucleotide antibiotic microcin C, a translation inhibitor that targets the enzyme aspartyl-tRNA synthetase. Furthermore, it was recently shown that the Opp transporter from P. aeruginosa PAO1, which is identical to Npp, is required for uptake of the uridyl peptide antibiotic pacidamycin, which targets the enzyme translocase I (MraY), which is involved in peptidoglycan synthesis. We used several approaches to further explore the substrate specificity of the Npp transporter. Assays of growth in defined minimal medium containing peptides of various lengths and amino acid compositions as sole nitrogen sources, as well as Biolog Phenotype MicroArrays, showed that the Npp transporter is not required for di-, tri-, and oligopeptide uptake. Overexpression of the npp operon increased susceptibility not just to pacidamycin but also to nickel chloride and the peptidyl nucleoside antibiotic blasticidin S. Furthermore, heterologous expression of the npp operon in a yej-deficient mutant of E. coli resulted in increased susceptibility to albomycin, a naturally occurring sideromycin with a peptidyl nucleoside antibiotic. Additionally, heterologous expression showed that microcin C is recognized by the P. aeruginosa Npp system. Overall, these results suggest that the NppA1A2BCD transporter is involved in the uptake of peptidyl nucleoside antibiotics by P. aeruginosa PA14. IMPORTANCEOne of the world's most serious health problems is the rise of antibiotic-resistant bacteria. There is a desperate need to find novel antibiotic therapeutics that either act on new biological targets or are able to bypass known resistance mechanisms. Bacterial ABC transporters play an important role in nutrient uptake from the environment. These uptake systems could also be exploited by a Trojan horse strategy to facilitate the transport of antibiotics into bacterial cells. Several natural antibiotics mimic substrates of peptide uptake routes. In this study, we analyzed an ABC transporter involved in the uptake of nucleoside peptidyl antibiotics. Our data might help to design drug conjugates that may hijack this uptake system to gain access to cells.

Selective substrate uptake: The role of ATP-binding cassette (ABC) importers in pathogenesis

“…These antibiotics mimic essential nutrients, such as iron-siderophore complexes or peptides, and are imported to the cell by ABC transporters [121]. A peptidyl nucleoside antibiotic, pacidamycin, is imported by the nppA1A2BCD system and inhibits translocase I, MraY, in Pseudomonas aeruginosa [122,123]. The E. coli yejABEF system imports microcin C, a peptide-nucleotide antibiotic, which blocks the synthesis of tRNA asp [124].…”

Bacterial peptide transporters: Messengers of nutrition to virulence

“…The ability of these proteins to transport peptides is demonstrated frequently. 13,25,26 Proton coupled peptide transporters…”

Deciphering the Function of New Gonococcal Vaccine Antigens Using Phenotypic Microarrays

“…To increase the number of conditions available for testing, bacterial PMs have been designed to comprehensively screen bacterial strains for unique phenotypes (18). These arrays have been used to assess the fitness cost of antimicrobial resistance (19), to determine whether the increased pathogenicity of sequence type 131 extraintestinal pathogenic Escherichia coli is due to an altered metabolic profile (20), to examine the biological role of two-component systems in E. coli (21), and to identify the physiological functions of genes (22,23). Here, we have optimized growth conditions (culture medium, inoculum size, and incubation time) for N. gonorrhoeae and used them for the first PM analysis of N. gonorrhoeae to assess the physiological functions of vaccine candidate proteins NGO1205, NGO2054, NGO2121, NGO1985, NGO2054, chromosome locus J]); however, in N. gonorrhoeae, it does not contain an invariant cysteine residue, a hallmark feature of all lipoproteins (4).…”