Binding of CDR‐derived peptides is mechanistically different from that of high‐affinity parental antibodies

Abstract

We present data that reveal crucial differences between the binding mode of anti-gastrin17 (G17, pyroEGPW-LEEEEEAYGWMDF-NH 2 ) monoclonal antibodies (mAbs) and their CDR-derived synthetic binders (SBs) with G17. The mAbs recognize the N-terminal sequence of G17 ( pyroEGPWL) with nanomolar affinity and high sequence selectivity. Molecular simulations suggest that G17 recognition is based primarily on a multitude of weak antibody-ligand interactions (H-bonding, van der Waals, etc.) inside a structurally well-defined cleft-like binding pocket. Relatively small structural changes (e.g. G-2 to A for G17) have a drastic impact on affinity, which is characteristic for antibody-like binding. In contrast, SBs recognize various sequences, including G17-unrelated targets with affinities of 1:1 complexes estimated in the 0.1-1.0 mM range. In most cases however, the G17/SB complex stoichiometries are not well-defined, giving rise to multimer aggregate formation with high apparent complex stabilities. Mutational studies on both G17 and SBs reveal the importance of positively charged (K/R) and aromatic residues (W/Y/F) for G17/SB complex formation. We propose that the synthetic binders use combinations of electrostatic, hydrophobic, and/or cation-p interactions in a variety of ways due to their intrinsic flexibility. This may also be the reason for their relatively low target specificity. We speculate that our findings are of general relevance, in showing that high-affinity mAbs do not necessarily provide the optimal basis for functional mimics design.

Synthetic Peptides as Protein Mimics

“…Nevertheless, neutralization of gastrin in cell-based assays by the mimetic peptides could be demonstrated (Timmerman et al, 2009 ). The mode of action of the peptides, however, may be different from that of the parent antibodies (Timmerman et al, 2010 ), leading to the conclusion that further efforts in peptide design have to be made.…”

Development of Next-Generation Peptide Binders Using In vitro Display Technologies and Their Potential Applications

“…It is worth noting that the selected bicyclic peptide exhibited high inhibitory activity with an inhibition constant ( K i ) of 1.5 nM, and efficiently interrupted the intrinsic coagulation pathway in human plasma ex vivo . Moreover, various chemical approaches for synthesizing structurally constrained peptides are reported to strengthen target affinities and provide adequate function (21–23). These results demonstrate that structurally constrained peptides have both increased target affinity and proteolytic stability and also suggest that they are potential new drugs with the advantages of small molecules and biologics.…”

Peptide Paratope Mimics of the Broadly Neutralizing HIV‐1 Antibody b12

“…The CDRs, ranging in length from seven to 25 amino acids, are sequentially nonconsecutive: that is, they can be considered discontinuous binding sites of the antibody for its antigen. Paratope mimetic peptides have been reported for a range of antibodies that recognize diverse antigens, including the HER2 receptor, CD4, and gastrin, as well as the V3 loop of HIV‐1 gp120 . These peptides present individual CDRs of their parent antibodies, with the exception of the anti‐CD4 antibody mimetic peptide, which presents key amino acid residues of all CDRs in one cyclic peptide.…”