Role of electrostatic interactions for ligand recognition and specificity of peptide transporters

Abstract

BackgroundPeptide transporters are membrane proteins that mediate the cellular uptake of di- and tripeptides, and of peptidomimetic drugs such as β-lactam antibiotics, antiviral drugs and antineoplastic agents. In spite of their high physiological and pharmaceutical importance, the molecular recognition by these transporters of the amino acid side chains of short peptides and thus the mechanisms for substrate binding and specificity are far from being understood.ResultsThe X-ray crystal structure of the peptide transporter YePEPT from the bacterium Yersinia enterocolitica together with functional studies have unveiled the molecular bases for recognition, binding and specificity of dipeptides with a charged amino acid residue at the N-terminal position. In wild-type YePEPT, the significant specificity for the dipeptides Asp-Ala and Glu-Ala is defined by electrostatic interaction between the in the structure identified positively charged Lys314 and the negatively charged amino acid side chain of these dipeptides. Mutagenesis of Lys314 into the negatively charged residue Glu allowed tuning of the substrate specificity of YePEPT for the positively charged dipeptide Lys-Ala. Importantly, molecular insights acquired from the prokaryotic peptide transporter YePEPT combined with mutagenesis and functional uptake studies with human PEPT1 expressed in Xenopus oocytes also allowed tuning of human PEPT1’s substrate specificity, thus improving our understanding of substrate recognition and specificity of this physiologically and pharmaceutically important peptide transporter.ConclusionThis study provides the molecular bases for recognition, binding and specificity of peptide transporters for dipeptides with a charged amino acid residue at the N-terminal position.Electronic supplementary materialThe online version of this article (doi:10.1186/s12915-015-0167-8) contains supplementary material, which is available to authorized users.

Tripeptide binding in a proton‐dependent oligopeptide transporter

“…During the transport cycle, these domains move relative to each other to allow alternate access from the cytoplasmic and extracellular sides . Several X‐ray structures of bacterial POTs have been reported, either in the apo‐form , peptide bound state , or in complex with the phosphonodipeptide alafosfalin (Table S1). However, only three structures have hitherto been reported in which a tripeptide is bound.…”

“…Several X-ray structures of bacterial POTs have been reported, either in the apo-form [8][9][10][11][12][13][14][15][16], peptide bound state [17][18][19][20], or in complex with the phosphonodipeptide alafosfalin [21,22] (Table S1). However, only three structures have hitherto been reported in which a tripeptide is bound.…”

An Optimized Screen Reduces the Number of GA Transporters and provides Insights into NPF Substrate Determinants

“…As cluster I and II contains the majority of GA transporters it is possible that some of these 568 residues are important for GA recognition. Incidentally, an investigation of YePepT from Yersinia 569 enterocolitica identified Lys314 as a determinant of specificity towards negatively charged 570 dipeptides (Boggavarapu et al, 2015). and that other specificities are known for members of clusters I and II, our analyses indicate that GA 577 may be recognized differently by different NPF GA transporters.…”

Peptide Selectivity of the Proton-Coupled Oligopeptide Transporter from <b><i>Neisseria meningitidis</i></b>

“…On the contrary YdgR and YhiP from E. coli show specificity to a much lesser extent [Harder et al, 2008;Weitz et al, 2007]. Other [Guettou et al, 2014] and YePEPT from Yersinia enterocolitica [Boggavarapu et al, 2015], which both have a preference for an acidic residue on the N-terminal residue of a dipeptide, and in the case of PepT So also in a tripeptide [Prabhala et al, 2014a]. For comparison with NmPOT, a more relevant example is DtpT from Lactococcus lactis ; in the case of this transporter the apparent affinity for dipeptides with a positively charged side chain in the N-terminal position decreases 80-fold compared to Ala-Ala, but no change is observed when a positively charged side chain is introduced in the N-terminus.…”