Single-molecule dynamics of the DNA-EcoRII protein complexes revealed with high-speed atomic force microscopy

Jamie L. Gilmore, Yuki Suzuki, Gintautas Tamulaitis, Virginijus Siksnys, Kunio Takeyasu, Yuri L Lyubchenko

Research output: Contribution to journalArticle

55 Citations (Scopus)

Abstract

The study of interactions of protein with DNA is important for gaining a fundamental understanding of how numerous biological processes occur, including recombination, transcription, repair, etc. In this study, we use the EcoRII restriction enzyme, which employs a three-site binding mechanism to catalyze cleavage of a single recognition site. Using high-speed atomic force microscopy (HS-AFM) to image single-molecule interactions in real time, we were able to observe binding, translocation, and dissociation mechanisms of the EcoRII protein. The results show that the protein can translocate along DNA to search for the specific binding site. Also, once specifically bound at a single site, the protein is capable of translocating along the DNA to locate the second specific binding site. Furthermore, two alternative modes of dissociation of the EcoRII protein from the loop structure were observed, which result in the protein stably bound as monomers to two sites or bound to a single site as a dimer. From these observations, we propose a model in which this pathway is involved in the formation and dynamics of a catalytically active three-site complex.

Original languageEnglish (US)
Pages (from-to)10492-10498
Number of pages7
JournalBiochemistry
Volume48
Issue number44
DOIs
StatePublished - Nov 10 2009

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Atomic Force Microscopy
Atomic force microscopy
Molecules
DNA
Proteins
Binding Sites
Biological Phenomena
Recombinational DNA Repair
Transcription
Dimers
Catalytic Domain
Repair
Monomers

ASJC Scopus subject areas

  • Biochemistry

Cite this

Single-molecule dynamics of the DNA-EcoRII protein complexes revealed with high-speed atomic force microscopy. / Gilmore, Jamie L.; Suzuki, Yuki; Tamulaitis, Gintautas; Siksnys, Virginijus; Takeyasu, Kunio; Lyubchenko, Yuri L.

In: Biochemistry, Vol. 48, No. 44, 10.11.2009, p. 10492-10498.

Research output: Contribution to journalArticle

Gilmore, Jamie L. ; Suzuki, Yuki ; Tamulaitis, Gintautas ; Siksnys, Virginijus ; Takeyasu, Kunio ; Lyubchenko, Yuri L. / Single-molecule dynamics of the DNA-EcoRII protein complexes revealed with high-speed atomic force microscopy. In: Biochemistry. 2009 ; Vol. 48, No. 44. pp. 10492-10498.
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