Quaternary Structure of SecA in Solution and Bound to SecYEG Probed at the Single Molecule Level

Ilja Kusters, Geert van den Bogaart, Alexej Kedrov, Victor Krasnikov, Faizah Fulyani, Bert Poolman, Arnold J. M. Driessen*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

48 Citations (Scopus)
286 Downloads (Pure)

Abstract

Dual-color fluorescence-burst analysis (DCFBA) was applied to measure the quaternary structure and high-affinity binding of the bacterial motor protein SecA to the protein-conducting channel SecYEG reconstituted into lipid vesicles. DCFBA is an equilibrium technique that enables the direct observation and quantification of protein-protein interactions at the single molecule level. SecA binds to SecYEG as a dimer with a nucleotide- and preprotein-dependent dissociation constant. One of the SecA protomers binds SecYEG in a salt-resistant manner, whereas binding of the second protomer is salt sensitive. Because protein translocation is salt sensitive, we conclude that the dimeric state of SecA is required for protein translocation. A structural model for the dimeric assembly of SecA while bound to SecYEG is proposed based on the crystal structures of the Thermotoga maritima SecA-SecYEG and the Escherichia coil SecA dinner.

Original languageEnglish
Pages (from-to)430-439
Number of pages10
JournalStructure
Volume19
Issue number3
DOIs
Publication statusPublished - 9-Mar-2011

Keywords

  • FLUORESCENCE-BURST ANALYSIS
  • PROTEIN-TRANSLOCATION CHANNEL
  • ESCHERICHIA-COLI
  • DIMERIC SECA
  • PREPROTEIN TRANSLOCATION
  • ATPASE SECA
  • CORRELATION SPECTROSCOPY
  • COINCIDENCE ANALYSIS
  • CRYSTAL-STRUCTURE
  • OLIGOMERIC STATE

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