Lynn Richardson

  • Jan 30, 2018

Date & Location: January 30, 2018, at 12p; Room 168 Plant Biology Building

Subject: Molecular topology of the transit peptide during chloroplast protein import

Abstract: Chloroplast protein import is directed by the interaction of the targeting signal (transit peptide) of nuclear encoded preproteins with translocons at the outer (TOC) and inner (TIC) chloroplast envelope membranes. Studies of the energetics and determinants of transit peptide binding to TOC and TIC have led to the hypothesis that import occurs through sequential recognition of transit peptides by components of TOC-TIC to facilitate preprotein recognition and membrane translocation.

To explore the molecular interactions at each stage of import, we employed a site-specific crosslinking approach to map the topology of the transit peptide in relation to TOC-TIC components by capturing specific stages in import. We demonstrate that the transit peptide penetrates the TOC complex and interacts with components of the TIC complex at early stages of binding in the absence of nucleoside triphosphate (NTP) hydrolysis. Low levels of ATP are required to stabilize the association of the preprotein with TOC-TIC, and GTP hydrolysis at the TOC receptors, Toc34 and Toc159, appears to play a previously unrecognized role in regulating release of the preprotein into the stroma. Tic20 is the major target of cross-linking at the inner membrane, supporting its central role in the TIC machinery. Cross-linking of the transit peptide at all stages of import occurs in large TOC-TIC supercomplexes, demonstrating the close cooperativity of the two membranes during all stages of protein import.