Peptide transporter DtpA has two alternate conformations, one of which is promoted by inhibitor binding

Bippes, Christian A.; Ge, Lin; Meury, Marcel; Harder, Daniel; Ucurum, Zöhre; Daniel, Hannelore; Fotiadis, Dimitrios; Müller, Daniel J. (2013). Peptide transporter DtpA has two alternate conformations, one of which is promoted by inhibitor binding. Proceedings of the National Academy of Sciences of the United States of America - PNAS, 110(42), E3978-E3986. National Academy of Sciences NAS 10.1073/pnas.1312959110

Full text not available from this repository. (Request a copy)

Peptide transporters (PTRs) of the large PTR family facilitate the uptake of di- and tripeptides to provide cells with amino acids for protein synthesis and for metabolic intermediates. Although several PTRs have been structurally and functionally characterized, how drugs modulate peptide transport remains unclear. To obtain insight into this mechanism, we characterize inhibitor binding to the Escherichia coli PTR dipeptide and tripeptide permease A (DtpA), which shows substrate specificities similar to its human homolog hPEPT1. After demonstrating that Lys[Z-NO2]-Val, the strongest inhibitor of hPEPT1, also acts as a high-affinity inhibitor for DtpA, we used single-molecule force spectroscopy to localize the structural segments stabilizing the peptide transporter and investigated which of these structural segments change stability upon inhibitor binding. This characterization was done with DtpA embedded in the lipid membrane and exposed to physiologically relevant conditions. In the unbound state, DtpA adopts two main alternate conformations in which transmembrane α-helix (TMH) 2 is either stabilized (in ∼43% of DtpA molecules) or not (in ∼57% of DtpA molecules). The two conformations are understood to represent the inward- and outward-facing conformational states of the transporter. With increasing inhibitor concentration, the conformation characterized by a stabilized TMH 2 becomes increasingly prevalent, reaching ∼92% at saturation. Our measurements further suggest that Lys[Z-NO2]-Val interacts with discrete residues in TMH 2 that are important for ligand binding and substrate affinity. These interactions in turn stabilize TMH 2, thereby promoting the inhibited conformation of DtpA.

Item Type:

Journal Article (Original Article)

Division/Institute:

04 Faculty of Medicine > Pre-clinic Human Medicine > Institute of Biochemistry and Molecular Medicine

UniBE Contributor:

Meury, Marcel; Harder, Daniel; Ucurum Fotiadis, Zöhre and Fotiadis, Dimitrios José

Subjects:

500 Science > 570 Life sciences; biology
600 Technology > 610 Medicine & health

ISSN:

0027-8424

Publisher:

National Academy of Sciences NAS

Submitter:

Patrizia Catucci

Date Deposited:

01 May 2014 12:47

Last Modified:

01 May 2014 12:47

Publisher DOI:

10.1073/pnas.1312959110

PubMed ID:

24082128

Uncontrolled Keywords:

atomic force microscopy major facilitator superfamily membrane transporter molecular interactions proton-dependent oligopeptide transporter

URI:

https://boris.unibe.ch/id/eprint/43403

Actions (login required)

Edit item Edit item
Provide Feedback