Kinetics and Consequences of Binding of Nona- and Dodecapeptides to the Oligopeptide Binding Protein (OppA) of Lactococcus lactis

Abstract

The oligopeptide transport system (Opp) of Lactococcus lactis belongs to the class of binding protein-dependent ABC-transporters. This system has the unique capacity to mediate the uptake of peptides from 4 up to at least 18 residues. Kinetic analysis of peptide binding to the binding protein, OppA, revealed a relationship between the peptide dissociation constants and the length of the ligand. The dissociation constants varied from submicromolar for dodecapeptides to millimolar for pentapeptides. This implies that the residues 6-12 of the peptide contribute to the binding affinity, and, in contrast to the current views on peptide binding by homologous proteins, these residues must interact with OppA. Analysis of pre-steady-state kinetics of binding showed that the observed differences in the -values result primarily from variations in the dissociation rate constants. These results are discussed in relation to the affinity constant for transport of these substrates. Overall, the data suggest that the slow dissociation rate constants for the larger peptides are rate determining in the translocation of peptides across the membrane.

The specificity of oligopeptide transport by Streptococcus thermophilus resembles that of Lactococcus lactis and not that of pathogenic streptococci

“…Nevertheless, peptide transport has been reported to limit the growth rate of the strain in media containing casein-derived peptides (Letort et al, 2002). It therefore suggests that this limitation results from kinetic considerations, presumably the rate of peptide donation from the binding protein(s) to the transmembrane complex (see below), as is the case for L. lactis (Lanfermeijer et al, 1999), rather than from a limited range of peptide preference.…”

Article

“…L'inactivation du gène oppA [16,26] a permis de mettre en évi-dence que Opp, chez MG1363, est le seul système impliqué dans le transport des oligopeptides supérieur à trois résidus, ceci malgré la présence dans le génome de cette souche d'autres opérons codant pour des transporteurs de peptides putatifs [3].…”

“…Le modèle de fonctionnement du système Opp, établi par Lanfermeijer et al [16] décompose le transport de peptides en quatre étapes : I, fixation réversible du ligand à la protéine affine OppA (dans sa forme ouverte) ; II, changement de conformation de la protéine OppA permettant la capture du peptide (forme OppA fermée) ; III, rapprochement de la protéine OppA liée au substrat vers le complexe perméase OppBC, changement de conformation du complexe perméase ; IV, translocation du substrat à travers la membrane (Fig. 1).…”

“…-Il existe une relation entre l'affinité pour le substrat et sa taille : plus le peptide est long, plus l'affinité OppA/peptide est importante avec une affinité maximale pour les nonapeptides [16]. La capacité de fixation peut s'étendre jusqu'à 35 résidus [8].…”

Peptides and ATP binding cassette peptide transporters

“…This suggestion is in line with the observation that the ABC transporter OpuA has two ligand binding domains per functional complex. Thirdly, depending on which step is ratedetermining in the translocation process, the K m for transport may or may not reflect the K d for binding [10,29]. Further work is needed to discriminate between these possibilities.…”