Molecular Diversity

, Volume 1, Issue 1, pp 29–38 | Cite as

Libraries of random-sequence polypeptides produced with high yield as carboxy-terminal fusions with ubiquitin

  • Thomas H. LaBean
  • Stuart A. Kauffman
  • Tauseef R. Butt
Research Papers

Summary

Libraries of random-sequence polypeptides have been shown to be valuable sources of novel molecules possessing a variety of useful biologic-like activities, some of which may hold promise as potential vaccines and therapeutics. Previous random peptide expression systems were limited to low levels of peptide production and often to short sequences. Here we describe a series of libraries designed for increased polypeptide length. Cloned as carboxy-terminal extensions of ubiquitin, the fusions were produced inE. coli at high levels, and were purified to homogeneity. The majority of the extension proteins examined could be cleaved from ubiquitin by treatment with a ubiquitin-fusion hydrolase. The libraries described here are appropriate sources of novel polypeptides with desired binding or catalytic function, as well as tools with which to examine inherent properties of proteins as a whole. Toward the latter goal, we have examined structural properties of random-sequence proteins purified from these libraries. Quite surprisingly, fluorescence emission spectra of intrinsic tryptophan residues in several purified fusion proteins, under native-like and denaturing conditions, often resemble those expected for folded and unfolded states, respectively. The results presented here detail an important expansion in the range of potential uses for random-sequence polypeptide libraries.

Keywords

Combinatorial library Random-sequence polypeptide Ubiquitin Ubiquitin fusion Protein folding 

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Copyright information

© ESCOM Science Publishers B.V. 1995

Authors and Affiliations

  • Thomas H. LaBean
    • 1
  • Stuart A. Kauffman
    • 1
    • 2
  • Tauseef R. Butt
    • 1
    • 3
  1. 1.Department of Biochemistry and BiophysicsUniversity of PennsylvaniaPhiladelphiaUSA
  2. 2.Santa Fe InstituteSanta FeUSA
  3. 3.Department of Molecular Virology and Host Defense, Research and DevelopmentSmithKline Beecham PharmaceuticalsKing of PrussiaUSA

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