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Protease protection assays show polypeptide movement into the SecY channel by power strokes of the SecA ATPase
Open accessAbstract
Bacterial secretory proteins are translocated post-translationally by the SecA ATPase through the protein-conducting SecY channel in the plasma membrane. During the ATP hydrolysis cycle, SecA undergoes large conformational changes of its two-helix finger and clamp domains, but how these changes result in polypeptide movement is unclear. Here, we use a reconstituted purified system and protease protection assays to show that ATP binding to SecA results in a segment of the translocation substrate being pushed into the channel. This motion is prevented by mutation of conserved residues at the finger's tip. Mutation of SecA's clamp causes backsliding of the substrate in the ATP-bound state. Together, these data support a power stroke model of translocation in which, upon ATP binding, the two-helix finger pushes the substrate into the channel, where it is held by the clamp until nucleotide hydrolysis has occurred.
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Translocation of integral membrane proteins was simulated at the cost of 1 ATP per 10 amino acids as observed in Catipovic and Rapoport (2020).
“Translocation of integral membrane proteins was simulated by a process in which each protein is translocated at the cost of 1 ATP per 10 amino acids as was observed in Catipovic and Rapoport (2020).”
From Fundamental behaviors emerge from simulations of a living minimal cell (2022) · cites this paper as Catipovic and Rapoport (2020)
The Results and Discussion (page unavailable) estimate that SecA may move about 10 amino acids per power stroke, but say the exact step size cannot be determined and varies with substrate sequence. The experiments used proOmpA, a model secretory protein; they did not establish a cost of one ATP per 10 amino acids for integral membrane proteins. (LLM, conf=0.88)
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References 21 · Cited by 11
Research works printed but not linked: 18
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Metadata
- Authors
- Catipovic, M. A., & Rapoport, T. A.
- Affiliations
- Howard Hughes Medical Institute (US)
- Corresponding author
- Rapoport, Tom A
- Venue
- The EMBO Reports
- Published
- 2020-09-24
- Volume
- 21
- Issue
- 11
- Pages
- e50905-e50905
- ISSN
- 1469-221X (ISSN-L)
- 1469-3178
- PMC
- PMC7645225
- OpenAlex
- W3088338476
- Cited by
- 11
- References
- 21
- Citations by year
- 2024: 2 · 2023: 3 · 2022: 4 · 2021: 2
- FWCI
- 1.30
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Locations
- EMBO Reports (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Funders
- NIGMS (R01GM052586)
- Howard Hughes Medical Institute
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › ATP Synthase and ATPases Research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Heat shock proteins research
- Keywords
- SecA ATPase, protein translocation, ATP binding, ATP hydrolysis, bacterial protein secretion, conformational change
- MeSH terms
- SEC Translocation Channels, SecA Proteins, Adenosine Triphosphatases, Bacterial Proteins, Models, Molecular, Peptide Hydrolases, Peptides, Protein Transport, Membrane Transport Proteins, Escherichia coli Proteins
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