Testing mechanisms of DNA sliding by architectural DNA-binding proteins: Dynamics of single wild-type and mutant protein molecules in vitro and in vivo

Kiyoto Kamagata, Yuji Itoh, Cheng Tan, Eriko Mano, Yining Wu, Sridhar Mandali, Shoji Takada, Reid C. Johnson

Research output: Contribution to journalArticlepeer-review

2 Citations (Scopus)

Abstract

Architectural DNA-binding proteins (ADBPs) are abundant constituents of eukaryotic or bacterial chromosomes that bind DNA promiscuously and function in diverse DNA reactions. They generate large conformational changes in DNA upon binding yet can slide along DNA when searching for functional binding sites. Here we investigate the mechanism by which ADBPs diffuse on DNA by single-molecule analyses of mutant proteins rationally chosen to distinguish between rotation-coupled diffusion and DNA surface sliding after transient unbinding from the groove(s). The properties of yeast Nhp6A mutant proteins, combined with molecular dynamics simulations, suggest Nhp6A switches between two binding modes: a static state, in which the HMGB domain is bound within the minor groove with the DNA highly bent, and a mobile state, where the protein is traveling along the DNA surface by means of its flexible N-terminal basic arm. The behaviors of Fis mutants, a bacterial nucleoid-associated helix-turn-helix dimer, are best explained by mobile proteins unbinding from the major groove and diffusing along the DNA surface. Nhp6A, Fis, and bacterial HU are all near exclusively associated with the chromosome, as packaged within the bacterial nucleoid, and can be modeled by three diffusion modes where HU exhibits the fastest and Fis the slowest diffusion.

Original languageEnglish
Pages (from-to)8642-8664
Number of pages23
JournalNucleic acids research
Volume49
Issue number15
DOIs
Publication statusPublished - 2021 Sep 7

ASJC Scopus subject areas

  • Genetics

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