Expanding the scope of CE reactor to ssDNA-binding protein-ssDNA complexes as exemplified for a tool for direct measurement of dissociation kinetics of biomolecular complexes

Toru Takahashi, Kei Ichirou Ohtsuka, Yoriyuki Tomiya, Nobuhiko Iki, Hitoshi Hoshino

Research output: Contribution to journalArticlepeer-review

1 Citation (Scopus)

Abstract

CE reactor (CER), which was developed as a tool for direct measurement of the dissociation kinetics of metal complexes, was successfully applied to the complexes of Escherichia coli ssDNA-binding protein (SSB) with ssDNA. The basic concept of CER is the application of CE separation process as a dissociation kinetic reactor for the complex, and the observation of the on-capillary dissociation reaction profile of the complex as the decrease of the peak height of the complex with increase of the migration time. The peak height of [SSB-ssDNA] decreases as the migration time increases since the degree of the decrease of [SSB-ssDNA] through the on-capillary dissociation reaction is proportional to the degree of the decrease of the peak height of [SSB-ssDNA]. The dissociation degreetime profiles for the complexes are quantitatively described by analyzing a set of electropherograms with different migration times. Dissociation rate constants of [SSB-ssDNA] consisting of 20-mer, 25-mer and 31-mer ssDNA were directly determined to be 3.99 x 10-4, 4.82 x 10-4 and 1.50 x 10-3/s, respectively. CER is a concise and effective tool for dissociation kinetic analysis of biomolecular complexes.

Original languageEnglish
Pages (from-to)3079-3085
Number of pages7
JournalElectrophoresis
Volume30
Issue number17
DOIs
Publication statusPublished - 2009 Sep 1

Keywords

  • Biomolecular complex
  • CE
  • Dissociation kinetic analysis

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry

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