The purified E. coli integral membrane protein is sufficient for reconstitution of SecA-dependent precursor protein translocation

The ATPase domain of SecA can form a tetramer in solution 1 1Edited by I. B. Holland

“…22 Recent scanning transmission electron microscopy (STEM) studies indicate that detergent-solubilized SecYEG exists as monomers and dimers, and that these are assembled by SecA-(AMP-PNP) to form a tetrameric complex (i.e. Sec(YEG) 4 ). 23 Furthermore, arrested translocation reactions were found to contain particles of mass 531 and 686 kDa, which are thought to comprise preprotein substrate, SecA dimer, and either two or four SecYEG heterotrimers.…”

Sec-dependent protein export and the involvement of the molecular chaperone SecB

“…Individ- ual fractions obtained during the purification were assayed by forming proteoliposomes through detergent removal and measuring their capacity to support, in the presence of proOmpA, the translocation ATPase activity of SecA. The 3 members of this complex were identified in a sodium dodecyl sulfate-polyacrylamide gel as band 1, SecY, and SecE (Brundage et al 1990). SecY spans the membrane 10 times and SecE spans the membrane 3 times (Akiyama and Ito 1989;Schatz et al 1989).…”

“…A trimeric complex was isolated from detergent extracts of inner membranes (Brundage et al 1990). Individ- ual fractions obtained during the purification were assayed by forming proteoliposomes through detergent removal and measuring their capacity to support, in the presence of proOmpA, the translocation ATPase activity of SecA.…”

Overexpression of phosphatidylglycerophosphate synthase restores protein translocation in asecG deletion mutant ofEscherichia coli at low temperature

“…Reconstitution of the protein translocation apparatus into proteoliposomes has clarified that SecA, SecE and SecY are absolutely required for translocation [5,6]. SecG has been shown to be required for efficient protein translocation [7][8][9].…”