DNA chain length dependence of formation and dynamics of hMutSalpha.hMutLalpha.heteroduplex complexes.

Published

Journal Article

Formation of a ternary complex between human MutSalpha, MutLalpha, and heteroduplex DNA has been demonstrated by surface plasmon resonance spectroscopy and electrophoretic gel shift methods. Formation of the hMutLalpha.hMutSalpha.heteroduplex complex requires a mismatch and ATP hydrolysis, and depends on DNA chain length. Ternary complex formation was supported by a 200-base pair G-T heteroduplex, a 100-base pair substrate was somewhat less effective, and a 41-base pair heteroduplex was inactive. As judged by surface plasmon resonance spectroscopy, ternary complexes produced with the 200-base pair G-T DNA contained approximately 0.8 mol of hMutLalpha/mol of heteroduplex-bound hMutSalpha. Although the steady-state levels of the hMutLalpha.hMutSalpha. heteroduplex were substantial, this complex was found to turn over, as judged by surface plasmon resonance spectroscopy and electrophoretic gel shift analysis. With the former method, the majority of the complexes dissociated rapidly upon termination of protein flow, and dissociation occurred in the latter case upon challenge with competitor DNA. However, ternary complex dissociation as monitored by gel shift assay was prevented if both ends of the heteroduplex were physically blocked with streptavidin.biotin complexes. This observation suggests that, like hMutSalpha, the hMutLalpha.hMutSalpha complex can migrate along the helix contour to dissociate at DNA ends.

Full Text

Duke Authors

Cited Authors

  • Blackwell, LJ; Wang, S; Modrich, P

Published Date

  • August 31, 2001

Published In

Volume / Issue

  • 276 / 35

Start / End Page

  • 33233 - 33240

PubMed ID

  • 11441019

Pubmed Central ID

  • 11441019

International Standard Serial Number (ISSN)

  • 0021-9258

Digital Object Identifier (DOI)

  • 10.1074/jbc.M105076200

Language

  • eng

Conference Location

  • United States