Dissection of bacteriophage λ site-specific recombination using synthetic peptide

Geoffrey Cassell, Martha Klemm, Clemencia Pinilla, Anca Segall

Research output: Contribution to journalArticlepeer-review

40 Scopus citations

Abstract

A wide variety of tools have been used to dissect biochemical pathways, inhibitors being chief among them. Combinatorial approaches have made the search for inhibitors much more efficient. We have applied such an approach to identify hexapeptides which inhibit different steps in a site-specific recombination reaction mediated by the bacteriophage λ integrase protein. Integrase's mechanism is still incompletely understood, in large part because several pathway intermediates remain hard to isolate. Integrase-catalyzed recombination is very efficient, but if blocked, it is highly reversible to substrates; this combination makes some intermediates exceedingly transient. We have used synthetic peptide combinatorial libraries to screen for hexapeptides that affect the recombination pathway at different stages, and have identified two families of peptides: One probably blocks DNA cleavage, the other may stabilize the Holliday junction intermediates. These peptides do not resemble parts of integrase or any of the other helper functions in the pathway. The deconvolution of hexapeptide libraries based both on inhibition of an enzymatic reaction as well as on accumulation of reaction intermediates is a novel approach to finding useful tools for dissecting a biochemical pathway. (C) 2000 Academic Press.

Original languageEnglish (US)
Pages (from-to)1193-1202
Number of pages10
JournalJournal of Molecular Biology
Volume299
Issue number5
DOIs
StatePublished - Jun 23 2000
Externally publishedYes

Keywords

  • Bacteriophage λ integrase
  • Combinatorial chemistry
  • Holliday junction
  • Inhibitory peptides
  • Tyrosine recombinase

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