Binding of SecA to the SecYEG Complex Accelerates the Rate of Nucleotide Exchange on SecA

Natale, Paolo; Swaving, Jelto; Does, Chris van der; Keyzer, Jeanine de; Driessen, Arnold J. M.

doi:10.1074/jbc.m312892200

Cited by 25 publications

(36 citation statements)

References 44 publications

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“…Strikingly, the SecYEG-bound SecA undergoes ATP-dependent conformational changes that are not observed for the soluble SecA. These data provide structural insight into the mechanism of the SecYEG-induced elevated nucleotide release kinetics at the SecA protein (38).…”

mentioning

confidence: 73%

“…E. coli SecA contains seven tryptophan residues that are located in distinct domains, (13,15,39) (Figure 1). Recently, we have shown that binding of SecA to SecYEG results in a major acceleration of the nucleotide exchange rate at NBF-I in conjunction with a decreased nucleotidebinding affinity (38). This implies that there must be a unique conformation of SecA while bound to SecYEG.…”

Section: Discussionmentioning

confidence: 99%

“…The reduced acrylamide accessibility of Trp 723 in the absence of ATP and Trp 701 in the presence of ATP suggest that binding to SecYEG shields the tryptophan residue from the quencher and positions it in a hydrophilic environment like for instance the water-filled channel inside SecYEG (31) or SecA (34). It should be emphasized that under the conditions employed the bound ATP is immediately hydrolyzed to ADP (38). Indeed, preliminary evidence indicates that ADP and ATP induce similar responses with SecA Trp 701 (unpublished data).…”

Section: Discussionmentioning

confidence: 99%

“…Tryptophan emission spectra from 320 to 450 nm were scanned at a rate of 1 nm/s using an excitation wavelength of 295 nm. Fluorescence spectra were corrected by substacting the buffer or a control containing the tryptophanless SecYEG proteoliposomes for the measurements with the soluble SecA and SecYEG-bound SecA, respectively (38).…”

Section: Methodsmentioning

confidence: 99%

“…All single tryptophan mutants of SecA were overexpressed in E. coli MM66 (48) and purified to homogeneity. The purified SecA mutant proteins were tested for in Vitro translocation of proOmpA into proteoliposomes reconstituted with the tryptophanless SecYEG complex (38). With all single tryptophan SecA proteins, translocation of proOmpA could be demonstrated ( Figure 2B).…”

Section: Functional Complementation Of the E Coli Mm66 Temperature-smentioning

confidence: 99%

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Conformational State of the SecYEG-Bound SecA Probed by Single Tryptophan Fluorescence Spectroscopy

et al. 2005

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The SecYEG complex is a membrane-embedded channel that permits the passage of precursor proteins (preproteins) across the inner membrane of Escherichia coli. SecA is a molecular motor that associates with the SecYEG pore and drives the stepwise translocation of preproteins across the membrane through multiple cycles of ATP binding and hydrolysis. We have investigated the conformational state of soluble and SecYEG-bound SecA using single tryptophan mutants of SecA. The fluorescence spectral properties of the single tryptophans of SecA and their accessibility to the quencher acrylamide demonstrate that SecA undergoes a conformational change that results in a more compact structure upon binding of ATP and binding to the SecYEG pore. In addition, SecYEG-bound SecA undergoes ATP-dependent conformational changes that are not observed for soluble SecA. These data support a model in which binding to the SecYEG channel has a major impact on the SecA conformation.

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mentioning

confidence: 73%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Functional Complementation Of the E Coli Mm66 Temperature-smentioning

confidence: 99%

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Conformational State of the SecYEG-Bound SecA Probed by Single Tryptophan Fluorescence Spectroscopy

et al. 2005

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Stabilization of SecA ATPase by the primary cytoplasmic salt of Escherichia coli

2019

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Much is known about the structure, function, and stability of the SecA motor ATPase that powers the secretion of periplasmic proteins across the inner membrane of Escherichia coli. Most studies of SecA are carried out in buffered sodium or potassium chloride salt solutions. However, the principal intracellular salt of E. coli is potassium glutamate (KGlu), which is known to stabilize folded proteins and protein‐nucleic acid complexes. Here we report that KGlu stabilizes SecA, including its dimeric state, and increases its ATPase activity, suggesting that SecA is likely fully folded, stable, and active in vivo at 37°C. Furthermore, KGlu also stabilizes a precursor form of the secreted maltose‐binding protein.

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The protein-conducting channel SecYEG

Veenendaal

Does

Driessen

2004

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

114

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In bacteria, the translocase mediates the translocation of proteins into or across the cytosolic membrane. It consists of a membrane embedded protein-conducting channel and a peripherally associated motor domain, the ATPase SecA. The channel is formed by SecYEG, a multimeric protein complex that assembles into oligomeric forms. The structure and subunit composition of this protein-conducting channel is evolutionary conserved and a similar system is found in the endoplasmic reticulum of eukaryotes and the cytoplasmic membrane of archaea. The ribosome and other membrane proteins can associate with the protein-conducting channel complex and affect its activity or functionality.

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Binding of SecA to the SecYEG Complex Accelerates the Rate of Nucleotide Exchange on SecA

Cited by 25 publications

References 44 publications

Conformational State of the SecYEG-Bound SecA Probed by Single Tryptophan Fluorescence Spectroscopy

Conformational State of the SecYEG-Bound SecA Probed by Single Tryptophan Fluorescence Spectroscopy

Stabilization of SecA ATPase by the primary cytoplasmic salt of Escherichia coli

The protein-conducting channel SecYEG

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