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Direct selection for a catalytic mechanism from combinatorial antibody libraries

Academic Article
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Overview

related to degree

  • Lo, Chih-Hung Larry, Ph.D. in Biology, Scripps Research 1990 - 1995

authors

  • Janda, Kim
  • Lo, Chih-Hung Larry
  • Li, T. Y.
  • Barbas III, Carlos
  • Wirsching, P.
  • Lerner, Richard

publication date

  • March 1994

journal

  • Proceedings of the National Academy of Sciences of the United States of America  Journal

abstract

  • Semisynthetic combinatorial antibody library methodology in the phage-display format was used to select for a cysteine residue in complementarity-determining regions. Libraries were panned with an alpha-phenethyl pyridyl disulfide that undergoes disulfide interchange. Out of 10 randomly picked clones, two contained an unpaired cysteine, one of which was studied. The antibody catalyzed the hydrolysis of the corresponding thioester where the electrophilic carbonyl occupies the three-dimensional space that was defined by the reactive sulfur atom during selection. The reaction operates by covalent catalysis. Although the steady-state rate enhancement relative to the activated thiol ester substrate is modest, hydrolysis of the acylated cysteine intermediate is remarkably efficient with a catalytic advantage of about four orders of magnitude. The results suggest that iterative mechanism-based selection procedures can recapitulate the enzymatic mechanisms refined through evolution.

subject areas

  • Amino Acid Sequence
  • Antibodies, Catalytic
  • Catalysis
  • Disulfides
  • Immunoglobulin Fab Fragments
  • Models, Genetic
  • Models, Molecular
  • Molecular Sequence Data
  • Selection, Genetic
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Identity

International Standard Serial Number (ISSN)

  • 0027-8424

Digital Object Identifier (DOI)

  • 10.1073/pnas.91.7.2532

PubMed ID

  • 8146149
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Additional Document Info

start page

  • 2532

end page

  • 2536

volume

  • 91

issue

  • 7

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