Enzyme-mediated spatial segregation on individual polymeric support beads: Application to generation and screening of encoded combinatorial libraries

Josef Vágner, George Barany, Kit Lam, Viktor Krchňák, Nikolai F. Sepetov, James A. Ostrem, Peter Štrop, Michal Lebl

Research output: Contribution to journalArticle

79 Scopus citations

Abstract

Proteolysis of short N(α)-protected peptide substrates bound to polyoxyethylene-polystyrene beads releases selectively free amino sites in the enzyme-accessible 'surface' area. The substantial majority of functional sites in the 'interior' of the polymeric support are not reached by the enzyme and remain uncleaved (protected). Subsequent synthesis with two classes of orthogonal protecting groups-N(α)-tert-butyloxycarbonyl (Boc) and N(α)-9-fluorenylmethyloxycarbonyl (Fmoc)-allows generation of two structures on the same bead. The surface structure is available for receptor interactions, whereas the corresponding interior structure is used for coding. Coding structures are usually readily sequenceable peptides. This 'shaving' methodology was illustrated by the preparation of a peptide- encoded model peptide combinatorial library containing 1.0 x 105 members at ≃6-fold degeneracy. From this single library, good ligands were selected for three different receptors: anti-β-endorphin antibody, streptavidin, and thrombin, and the binding structures were deduced correctly by sequencing the coding peptides present on the same beads.

Original languageEnglish (US)
Pages (from-to)8194-8199
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume93
Issue number16
DOIs
StatePublished - Aug 6 1996
Externally publishedYes

Keywords

  • combinatorial chemistry
  • encoded libraries
  • peptides
  • proteolytic enzymes
  • solid-phase synthesis

ASJC Scopus subject areas

  • Genetics
  • General

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