SecYEG and SecA Are the Stoichiometric Components of Preprotein Translocase

Douville, Karen; Price, Albert; Eichler, Jerry; Economou, Anastassios; Wickner, William

doi:10.1074/jbc.270.34.20106

Cited by 128 publications

(174 citation statements)

References 62 publications

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“…Moreover, the prokaryotic nascent chain passes through only a single membrane in contrast to the eukaryotic secretory proteins that are exported to different organelles. For Sec A-mediated targeting, the secretory proteins (that can be translocated after release from the ribosome) are held in a competent state by interaction with Sec B and/or Sec A [both of which are components of the secretory system that includes also Sec Y, Sec E, Sec D, and Sec F (Douville et al, 1995)]. The role of the proposed translational pauses in the secretory proteins may allow interaction of the nascent peptide with Sec A or Sec B, which may be optimal in the presence of a domain.…”

Section: Cytosolic Versus Periplasmic Proteinsmentioning

confidence: 99%

Ribosome‐mediated translational pause and protein domain organization

1996

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Because regions on the messenger ribonucleic acid differ in the rate at which they are translated by the ribosome and because proteins can fold cotranslationally on the ribosome, a question arises as to whether the kinetics of translation influence the folding events in the growing nascent polypeptide chain. Translationally slow regions were identified on mRNAs for a set of 37 multidomain proteins from Escherichia coli with known three-dimensional structures. The frequencies of individual codons in mRNAs of highly expressed genes from E. coli were taken as a measure of codon translation speed. Analysis of codon usage in slow regions showed a consistency with the experimentally determined translation rates of codons; abundant codons that are translated with faster speeds compared with their synonymous codons were found to be avoided; rare codons that are translated at an unexpectedly higher rate were also found to be avoided in slow regions. The statistical significance of the occurrence of such slow regions on mRNA spans corresponding to the oligopeptide domain termini and linking regions on the encoded proteins was assessed. The amino acid type and the solvent accessibility of the residues coded by such slow regions were also examined. The results indicated that protein domain boundaries that mark higher-order structural organization are largely coded by translationally slow regions on the RNA and are composed of such amino acids that are stickier to the ribosome channel through which the synthesized polypeptide chain emerges into the cytoplasm. The translationally slow nucleotide regions on mRNA possess the potential to form hairpin secondary structures and such structures could further slow the movement of ribosome. The results point to an intriguing correlation between protein synthesis machinery and in vivo protein folding. Examination of available mutagenic data indicated that the effects of some of the reported mutations were consistent with our hypothesis.Keywords: codon usage; domain linkers; peptide channel; protein folding; ribosome; translation The rate of protein translation is nonuniform along the mRNA (Varenne et al., 1984). Ribosomes seem to pause as well as stack at specific regions on mRNA (Chaney & Morris, 1979;Wolin &Walter, 1988;Kim et al., 1991;Kim & Hollingsworth, 1992). Two of the mRNA features that slow the ribosome during translation are the presence of slow codons (Varenne et al., 1984;Bonekamp et al., 1985;Wolin & Walter, 1988) and the formation of higher order nucleotide structures (Chaney & Morris, 1979;Tu et al., 1992). The resultant translational pause may temporally separate the adjacent regions on the growing nascent polypeptide. Given that proteins can fold cotranslationally with the possible involvement of the ribosome (Phillips, 1966;Baldwin, 1975;Yonath, 1992;Kudlicki et al., 1994;Wiedmann et al., 1994;Brimacombe, 1995), the temporal separation might exert a phenotypic effect on the structural organization of the encoded protein. We show that as much as 70% of the do...

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Section: Cytosolic Versus Periplasmic Proteinsmentioning

confidence: 99%

Ribosome‐mediated translational pause and protein domain organization

1996

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“…Iodinated SecA has been used to analyse the conformational changes of SecA during preprotein translocation [21,27]. In the presence of preprotein and ATE or with the nonhydrolysable ATP analogue AMP-PNP alone, SecYEG-bound SecA undergoes a conformational change yielding a stable 30kDa fragment after proteolysis [21,27,28]. This fragment corresponds to a carboxy-terminal region of SecA [29] and its formation is reversed upon hydrolysis of ATE whereas it is stabilized by SecD and SecF [21].…”

Section: Topology and Membrane Insertion Of Seca Domainsmentioning

confidence: 99%

The Sec system

Driessen

Fekkes

Wolk

1998

Current Opinion in Microbiology

159

111

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The Sec system Driessen, A.J.M.; Fekkes, P.; van der Wolk, J.P.W. Take-down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim.Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons the number of authors shown on this cover page is limited to 10 maximum. Proteins designated to be secreted by Escherichia coil are synthesized with an amino-terminal signal peptide and associate as nascent chains with the export-specific chaperone SecB. Translocation occurs at a multisubunit membrane-bound enzyme termed translocase, which consists of a peripheral preprotein-binding site and an ATPase domain termed SecA, a core heterotrimeric integral membrane protein complex with SecY, SecE and SecG as subunits, and an accessory integral membrane protein complex containing SecD and SecE Major new insights have been gained into the cascade of preprotein targeting events and the enzymatic mechanism of preprotein translocation. It has become clear that preproteins are translocated in a stepwise fashion involving large nucleotide-induced conformational changes of the molecular motor SecA that propels the translocation reaction. Addresses

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“…However, these sequences were too short to be considered full-length and lacked the hydrophobic core region for SecA interaction (Wang et al, 2000) and the c-region carrying the cleavage site for the signal peptidase (von Heijne, 1998), which would minimally require the translocation pathway. The minimal Sec pathway translocation pore passes proteins through the SecYEG membraneembedded translocon (Fekkes and Driessen, 1999), which also has the minimal stoichiometric subunits of translocase for the integral membrane trimer (Douville et al, 1995). Thus, we hypothesized that the alkaline soluble expression curve corresponded to the Sec pathway containing the Sec translocon (Fig.…”

Section: Resultsmentioning

confidence: 99%

Novel GFP Expression Using a Short N-Terminal Polypeptide through the Defined Twin-Arginine Translocation (Tat) Pathway

Lee

Han

Kim

et al. 2011

Molecules and Cells

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Escherichia coli is frequently used as a convenient host organism for soluble recombinant protein expression. However, additional strategies are needed for proteins with complex folding characteristics. Here, we suggested that the acidic, neutral, and alkaline isoelectric point (pI) range curves correspond to the channels of the E. coli type-II cytoplasmic membrane translocation (periplasmic translocation) pathways of twin-arginine translocation (Tat), Yid, and general secretory pathway (Sec), respectively, for unfolded and folded target proteins by examining the characteristic pI values of the N-termini of the signal sequences or the leader sequences, matching with the known diameter of the translocation channels, and analyzing the N-terminal pI value of the signal sequences of the Tat substrates. To confirm these proposed translocation pathways, we investigated the soluble expression of the folded green fluorescent protein (GFP) with short N-terminal polypeptides exhibiting pI and hydrophilicity separately or collectively. This, in turn, revealed the existence of an anchor function with a specific directionality based on the N-terminal pI value (termed as N-terminal pI-specific directionality) and distinguished the presence of the E. coli type-II cytoplasmic membrane translocation pathways of Tat, Yid, and Sec for the unfolded and folded target proteins. We concluded that the pI value and hydrophilicity of the short N-terminal polypeptide, and the total translational efficiency of the target proteins based on the ΔG RNA value of the N-terminal coding regions are important factors for promoting more efficient translocation (secretion) through the largest diameter of the Tat channel. These results show that the short N-terminal polypeptide could substitute for the Tat signal sequence with improved efficiency.

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SecYEG and SecA Are the Stoichiometric Components of Preprotein Translocase

Cited by 128 publications

References 62 publications

Ribosome‐mediated translational pause and protein domain organization

Ribosome‐mediated translational pause and protein domain organization

The Sec system

Novel GFP Expression Using a Short N-Terminal Polypeptide through the Defined Twin-Arginine Translocation (Tat) Pathway

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