Exploring Non-obvious Hydrophobic Binding Pockets on Protein Surfaces: Increasing Affinities in Peptide-Protein Interactions.

A 42-residue polypeptide conjugated to a small-molecule organic ligand capable of targeting the phosphorylated side chain of Ser15 was shown to bind glycogen phosphorylase a (GPa) with a K value of 280 nm. The replacement of hydrophobic amino acids by Ala reduced affinities, whereas the incorporation of l-2-aminooctanoic acid (Aoc) increased them. Replacing Nle5, Ile9 and Leu12 by Aoc reduced the K value from 280 to 27 nm.

Conjugation of a Dipicolyl Chelate to Polypeptide Conjugates Increases Binding Affinities for Human Serum Albumin and Survival Times in Human Serum.

The affinity for human serum albumin (HSA) of a series of 2-5 kDa peptides covalently linked to 3,5-bis[[bis(2-pyridylmethyl)amino]methyl]benzoic acid, a dipicolyl chelator with micromolar affinity for Zn , was found by surface plasmon resonance to increase in the presence of 1 μm ZnCl at physiological pH. The dependence on polypeptide hydrophobicity was found to be minor, thus suggesting that the conjugates bound to the metal-binding site and not to the fatty-acid-binding site. The affinity of the conjugates increased strongly with the positive charge of the polypeptides, thus implicating the negatively charged protein surface surrounding the metal-binding site.