A conformationally homogeneous combinatorial peptide library.
In search for a rational way to convert the information encoded in peptide structures into peptidomimetics, major progress could be made by coupling the power of selection methods, now enormously increased in number as a result of the development of combinatorial peptide libraries, with the rational design of structure-inducing templates for the selectable sequences. The availability of libraries of peptides with predetermined structure would enable selection-driven peptidomimetic design, whereby a conformational model for the peptide pharmacophore would be directly derived from the screening, allowing the design of a suitable non-peptidic scaffold to replace the peptide backbone. We describe here the first example of a conformationally homogeneous combinatorial peptide library, which yields ligands with the expected structure upon selection. The library was built by randomising five positions in the alpha-helical portion of a 26 amino acid Cys2His2 consensus "zinc-finger" motif. Since in zinc-fingers metal coordination and folding are coupled, in our library metal-dependent binding represents a built-in control against the selection of structurally undefined sequences. The alpha-helical library was produced as both fusion with the pVIII protein of filamentous phage and soluble peptides by chemical synthesis, the latter enabling the expansion of the selectable repertoire by the inclusion of non-coded amino acids. The two libraries were independently screened with the same receptor (a monoclonal IgA reactive against the lipopolysaccharide of the human pathogen Shigella flexneri), yielding a very similar consensus. In particular, the peptides defined by both methods showed very strong, zinc-dependent binding to the IgA. The geometrical arrangement of the side-chains of the selected peptide pharmacophore was shown by circular dichroism, Co(II)-complex absorption and high-resolution NMR to be structurally invariant with respect to the parent zinc-finger.
Bianchi
E
E
Department of Biotechnology, Istituto di Ricerche di Biologia Molecolare, P. Angeletti (IRBM), Rome, Italy.
Folgori
A
A
Wallace
A
A
Nicotra
M
M
Acali
S
S
Phalipon
A
A
Barbato
G
G
Bazzo
R
R
Cortese
R
R
Felici
F
F
D000595:Amino Acid Sequence; D001483:Base Sequence; D002942:Circular Dichroism; D016384:Consensus Sequence; D015723:Gene Library; D007070:Immunoglobulin A / Q000378:metabolism; D008070:Lipopolysaccharides / Q000276:immunology; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins; D012641:Selection, Genetic; D012763:Shigella flexneri / Q000276:immunology; D013329:Structure-Activity Relationship; D015032:Zinc / Q000378:metabolism; D016335:Zinc Fingers / Q000235:genetics*
D007070:Immunoglobulin A; D008070:Lipopolysaccharides; D010455:Peptides; D011993:Recombinant Fusion Proteins; D015032:Zinc