The SecA and SecY subunits of translocase are the nearest neighbors of a translocating preprotein, shielding it from phospholipids.

Joly, John C.; Wickner, William

doi:10.1002/j.1460-2075.1993.tb05651.x

Cited by 176 publications

(146 citation statements)

References 74 publications

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“…The process of intraorganellar protein transport from the stroma into the thylakoids may thus resemble that of intracellular protein transport in ancestral cyanobacteria, whereas the process of protein transport from the cytosol into the stroma may represent that acquired after the endosymbiotic event . It was shown that translocation of a dihydrofolate reductase fusion protein across the Escherichia coli cytoplasmic membrane was blocked by methotrexate and NADPH (Joly and Wickner, 1993). This may be consistent from an evolutionary point of view with our findings that the thylakoids do not have the capacity to take up the methotrexate-bound, tightly folded dihydrofolate reductase fusion protein.…”

Section: Discussionsupporting

confidence: 92%

Chloroplast Protein Import

Endo

Kawakami

Goto

et al. 1994

European Journal of Biochemistry

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Proteins have to be at least partially unfolded upon passage through the biological membranes. Previous studies with a dihydrofolate reductase fusion protein containing a chloroplast transit peptide showed that stabilization of the tertiary structure of the fusion protein by binding of a ligand, methotrexate, failed to block its translocation across the envelopes, suggesting that chloroplast envelopes have strong activity to unfold proteins [America, T., Hageman, J., Guéra, A., Rook, F., Archer, K., Keegstra, K. & Weisbeek, P. (1994) Plant Mol. Biol. 24, 283–294]. In the present study, we have analyzed in vitro translocation of a fusion protein consisting of the entire plastocyanin precursor and dihydrofolate reductase across the chloroplast envelope membranes and the thylakoid membrane. In the presence of methotrexate, the fusion protein was imported into the stroma but its translocation across the thylakoid membrane was blocked. The fusion protein that bound to the envelope became susceptible to digestion by thermolysin. These results suggest that, while the envelope membranes can unfold the methotrexate‐bound fusion protein to allow its passage, the thylakoid membrane cannot unfold the fusion protein that has re‐bound to methotrexate in the stroma.

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Section: Discussionsupporting

confidence: 92%

Chloroplast Protein Import

Endo

Kawakami

Goto

et al. 1994

European Journal of Biochemistry

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“…Due to its location at the core of the transport machinery, it has been the focus of considerable research aimed at understanding its structure and function. Previous studies have shown that E. coli SecY comes into close contact with the translocating polypeptide chain (33) and, in yeast, the signal peptide forms a helix in the process (34). We have now shown that the interaction is saturable and specific for functional signal peptides; neither a nonfunctional signal peptide nor an unrelated peptide effectively competes for binding.…”

Section: Discussionmentioning

confidence: 67%

Demonstration of a Specific Escherichia coli SecY−Signal Peptide Interaction

et al. 2004

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Protein translocation in Escherichia coli is initiated by the interaction of a preprotein with the membrane translocase composed of a motor protein, SecA ATPase, and a membrane-embedded channel, the SecYEG complex. The extent to which the signal peptide region of the preprotein plays a role in SecYEG interactions is unclear, in part because studies in this area typically employ the entire preprotein. Using a synthetic signal peptide harboring a photoaffinity label in its hydrophobic core, we examined this interaction with SecYEG in a detergent micellar environment. The signal peptide was found to specifically bind SecY in a saturable manner and at levels comparable to those that stimulate SecA ATPase activity. Chemical and proteolytic cleavage of cross-linked SecY and analysis of the signal peptide adducts indicate that the binding was primarily to regions of the protein containing transmembrane domains seven and two. The signal peptide-SecY interaction was affected by the presence of SecA and nucleotides in a manner consistent with the transfer of signal peptide to SecY upon nucleotide hydrolysis at SecA. † This research was supported in part by National Institutes of Health Grant GM37639 (to D.A.K. Protein transport across, or integration into, biological membranes is a vital cellular process (1-3). Components of the Sec translocon, the membrane pore through which presecretory proteins (or membrane proteins) achieve membrane translocation (or integration), are the most conserved transport constituents throughout the three kingdoms of life (4).In Escherichia coli, the essential components of the translocase (5) include the membraneassociated form of SecA (6, 7) and the polytopic membrane proteins SecY, SecE (homologues of the mammalian Sec61α, Sec61γ, and the yeast ER 1 Sec61p, Sss1p, respectively), and SecG (8, 9); the latter three proteins form a stable trimeric SecYEG complex (10). SecA is an ATPase that powers the membrane translocation of hydrophilic polypeptides by coupling ATP hydrolysis with protein movement via concomitant SecA membrane insertion and deinsertion cycles (11,12). SecY protein has 10 transmembrane (TM1-TM10), six cytosolic (C1-C6), and five periplasmic (P1-P5) domains (13), and it forms the core of the passageway for the translocating polypeptide chain (14 (35,36). Suppressor analysis of prlA mutations using dysfunctional LamB signal peptides revealed that the suppressor mutations clustered in distinct regions, and TM7 of SecY was proposed to function in signal sequence recognition (37). Yet no investigation has focused on the direct interaction between a signal peptide and the E. coli translocon. Consequently, it remains unclear whether the nascent polypeptide chain is merely translocating in close proximity to SecY, SecY performs simply a proofreading function (37), or SecY is more intimately involved in the specific recognition of the signal peptide. Wang et al. Page 2Biochemistry. Author manuscript; available in PMC 2011 April 29. NIH-PA Author ManuscriptNIH-PA Author Manuscr...

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“…1,2 In bacteria, a central role in both processes is fulfilled by the integral membrane complex SecYEG, that forms the protein conducting channel within the cytoplasmic membrane. 3,4 Two different cytoplasmic partners can bind to SecYEG to induce opening of the channel and to provide the driving force for the translocation process. Membrane proteins are mostly inserted cotranslationally by SecYEG-bound ribosomes whereas secretory proteins and large extracellular domains of integral membrane proteins are translocated posttranslationally by the motor protein SecA.…”

Section: Introductionmentioning

confidence: 99%