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. 2005 Oct 26;127(42):14584-5.
doi: 10.1021/ja055050o.

A rapid library screen for tailoring beta-peptide structure and function

Joshua A Kritzer  1 , Nathan W Luedtke, Elizabeth A Harker, Alanna Schepartz
Affiliations

A rapid library screen for tailoring beta-peptide structure and function

Joshua A Kritzer et al. J Am Chem Soc. .

Abstract

Recently we described a beta-decapeptide (beta53-1) that folds into a 14-helix in aqueous solution, binds the oncoprotein hDM2 with submicromolar affinity, and inhibits the interaction of hDM2 with a peptide derived from the activation domain of p53 (p53AD). The solution structure of beta53-1 in CD3OH revealed an unexpected C-terminal unwinding that staggers the side chains comprising the hDM2 recognition epitope to better mimic those of p53AD. The structure-function relationship implied by this distortion suggested that a library of beta53-1 analogues possessing diversity along a nonrecognition face might contain molecules possessing greater affinity for hDM2. Here we describe (1) beta-peptide synthesis protocols that produce high quality one-bead-one-beta-peptide libraries suitable for on-bead screening without purification, (2) a versatile, scalable on-bead screen, and (3) a simple tandem mass spectrometry (MS/MS) decoding method. Using this procedure, we identified beta53-1 analogues with improved structural and functional properties.

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Figures

Figure 1
Figure 1
Helical net representations of (A) βNEG and (B) the 1000-member library, β53-1, and β53-8–10. β3X denotes a β3-homoamino acid; X is the common one-letter code for the analogous α-amino acid. Side chains on the recognition, variable, and salt-bridge faces are colored green, orange, and blue/red, respectively. IC50 values are derived from curve fits to the p53AD·hDM2 inhibition data in Figure 3A.
Figure 2
Figure 2
Representative fluorescence micrographs of OBOβ screen. (A) Mixture of β53-1 and βNEG beads treated with 500 nM BiohDM2 and 5 nM QDots-SA605 in BW buffer. (B) Beads from the 1000-member library treated as in (A) in BW+ buffer or (C) with 200 nM BiohDM2 and 5 nM QDots-SA605 in BW+ buffer and washed extensively.
Figure 3
Figure 3
(A) Fluorescence polarization analysis of the inhibition of p53AD15–31Flu·hDM21–188 complexation by p53AD15–31 (red), β53-1 (gray), β53-8 (blue), β53-9 (green), or β53-10 (purple). (B,C) Superposition of the solution structures of β53-1 (gray) and β53-8 (blue) highlighting the varied nonrecognition face (B) and the recognition face (C).
See this image and copyright information in PMC

References

    1. Kritzer JA, Lear JD, Hodsdon ME, Schepartz A. J Am Chem Soc. 2004;126:9468–9469. - PubMed
    1. Kritzer JA, Hodsdon ME, Schepartz A. J Am Chem Soc. 2005;127:4118–4119. - PMC - PubMed
    1. Kussie PH, Gorina S, Marechal V, Elenbaas B, Moreau J, Levine AJ, Pavletich NP. Science. 1996;274:948–953. - PubMed

    1. One-bead-one-compound libraries have been used extensively for ligand discovery using α-peptides, peptidomimetics, and some classes of small molecules. For reviews, see: Lam KS, Lebl M, Krchnak V. Chem Rev. 1997;97:411–448.Lam KS, Lehman AL, Song AM, Doan N, Enstrom AM, Maxwell J, Liu RW. Methods Enzymol. 2003;369:298–322.

    1. Murray JK, Farooqi B, Sadowsky JD, Scalf M, Freund WA, Smith LM, Chen J, Gellman SH. J Am Chem Soc. 2005;127:13271–13280. - PubMed

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