Involvement of a bifunctional, paired-like DNA-binding domain and a transpositional enhancer in Sleeping Beauty transposition

J Biol Chem. 2002 Sep 13;277(37):34581-8. doi: 10.1074/jbc.M204001200. Epub 2002 Jun 24.

Abstract

Sleeping Beauty (SB) is the most active Tc1/mariner-like transposon in vertebrate species. Each of the terminal inverted repeats (IRs) of SB contains two transposase-binding sites (DRs). This feature, termed the IR/DR structure, is conserved in a group of Tc1-like transposons. The DNA-binding region of SB transposase, similar to the paired domain of Pax proteins, consists of two helix-turn-helix subdomains (PAI + RED = PAIRED). The N-terminal PAI subdomain was found to play a dominant role in contacting the DRs. Transposase was able to bind to mutant sites retaining the 3' part of the DRs; thus, primary DNA binding is not sufficient to determine the specificity of the transposition reaction. The PAI subdomain was also found to bind to a transpositional enhancer-like sequence within the left IR of SB, and to mediate protein-protein interactions between transposase subunits. A tetrameric form of the transposase was detected in solution, consistent with an interaction between the IR/DR structure and a transposase tetramer. We propose a model in which the transpositional enhancer and the PAI subdomain stabilize complexes formed by a transposase tetramer bound at the IR/DR. These interactions may result in enhanced stability of synaptic complexes, which might explain the efficient transposition of Sleeping Beauty in vertebrate cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • DNA / metabolism*
  • DNA Transposable Elements / physiology*
  • DNA-Binding Proteins / chemistry
  • Dimerization
  • Enhancer Elements, Genetic / physiology*
  • HeLa Cells
  • Humans
  • Transposases / chemistry

Substances

  • DNA Transposable Elements
  • DNA-Binding Proteins
  • DNA
  • Tc1 transposase
  • Transposases