title: Biophysical characterization of the proton-coupled oligopeptide transporter YjdL.
abstract: Proton-coupled oligopeptide transporters (POTs) utilize the electrochemical proton gradient to facilitate uptake of di- or tripeptide molecules. YjdL is one of four POTs found in Escherichia coli. It has shown an extraordinary preference for di- rather than tripeptides, and is therefore significantly different from prototypical POTs such as the human hPepT1. Nonetheless YjdL contains several highly conserved POT residues, which include Glu388 that is located in the putative substrate binding cavity. Here we present biophysical characterization of WT-YjdL and Glu388Gln. Isothermal titration calorimetrical studies exhibit a K(d) of 14 μM for binding of Ala-Lys to WT-YjdL. Expectedly, no binding could be detected for the tripeptide Ala-Ala-Lys. Surprisingly however, binding could not be detected for Ala-Gln, although earlier studies indicated inhibitory potencies of Ala-Gln to be comparable to Ala-Lys (IC(50) values of 0.6 compared to 0.3mM). Finally, Ala-Lys binding to Glu388Gln was also undetectable which may support a previously suggested role in interaction with the ligand peptide N-terminus.
authors.affiliation: Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Johanne Mørch
authors.initials: JM
authors.lastname: Jensen
authors.forename: Fie C
authors.initials: FC
authors.lastname: Simonsen
authors.forename: Amir
authors.initials: A
authors.lastname: Mastali
authors.forename: Helle
authors.initials: H
authors.lastname: Hald
authors.forename: Ida
authors.initials: I
authors.lastname: Lillebro
authors.forename: Frederik
authors.initials: F
authors.lastname: Diness
authors.forename: Lars
authors.initials: L
authors.lastname: Olsen
authors.forename: Osman
authors.initials: O
authors.lastname: Mirza
mesh_terms: D001703:Biophysics; D002151:Calorimetry / Q000379:methods; D002850:Chromatography, Gel; D002942:Circular Dichroism; D004151:Dipeptides / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018698:Glutamic Acid / Q000737:chemistry; D005973:Glutamine / Q000737:chemistry; D008024:Ligands; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011522:Protons; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification
chemical_list: D004151:Dipeptides; D029968:Escherichia coli Proteins; D008024:Ligands; D026901:Membrane Transport Proteins; D011522:Protons; D011994:Recombinant Proteins; C544083:YjdL protein, E coli; D005973:Glutamine; D018698:Glutamic Acid