PEPTIDE BINDING IN OPPA, THE CRYSTAL-STRUCTURES OF THE PERIPLASMIC OLIGOPEPTIDE BINDING-PROTEIN IN THE UNLIGANDED FORM AND IN COMPLEX WITH LYSYLLYSINE

The periplasmic oligopeptide binding protein, OppA, acts as the initia l receptor for the uptake of peptides by the oligopeptide permease (Op p) in Gram-negative bacteria. Opp will handle peptides between two and five amino acid residues regardless of their sequence. The crystal st ructures of a series of OppA-peptide complexes have revealed an enclos ed but versatile peptide binding pocket and have illustrated how tri-a nd tetrapeptide ligands are accommodated. Here, the crystal structures of (i) OppA complexed with a dipeptide (lysyllysine) and (ii) unligan ded OppA have been solved using X-ray data extending to 1.8 and 2.4 An gstrom spacing, respectively. In the dipeptide complex, the alpha-amin o group of the ligand is anchored through an ion pair interaction with Asp(419), as observed in complexes with longer peptides. However, its alpha-carboxylate group forms water-mediated interactions with the gu anidinium groups of Arg(404) and Arg(413) rather than the direct salt bridges to Arg(413) and His(371) observed in the tripeptide and tetrap eptide complexes, respectively. Isothermal titration calorimetric meas urements of the binding of lysine-containing peptides of different len gths to OppA show that the dipeptide, KK, is bound with similar to 60- fold lower affinity than related tri-and tetrapeptides (KKK and KKKA, respectively). These data are discussed with reference to the calculat ed enthalpic and entropic contributions to ligand binding and the stru ctures of the OppA peptide complexes. In the unliganded molecule, doma in III has rotated as a rigid body through 26 degrees away from domain s I and II, exposing the ligand binding site. The water structure in t he binding cleft shows similarities to that in the various OppA-peptid e complexes.