The C-terminal tail of the bacterial translocation ATPase SecA modulates its activity
- University of Birmingham, United Kingdom
- Heidelberg University, Germany
- University of Heidelberg, Germany
Abstract
In bacteria, the translocation of proteins across the cytoplasmic membrane by the Sec machinery requires the ATPase SecA. SecA can bind ribosomes and recognise nascent substrate proteins, but the molecular mechanism of recognition is unknown. We investigated the role of the C-terminal tail (CTT) of SecA in nascent polypeptide recognition. The CTT consists of a flexible linker (FLD) and a small metal-binding domain (MBD). Phylogenetic analysis and ribosome binding experiments indicated that the MBD interacts with 70S ribosomes. Disruption of the MBD only or the entire CTT had opposing effects on ribosome binding, substrate-protein binding, ATPase activity and in vivo function, suggesting that the CTT influences the conformation of SecA. Site-specific crosslinking indicated that F399 in SecA contacts ribosomal protein uL29, and binding to nascent chains disrupts this interaction. Structural studies provided insight into the CTT-mediated conformational changes in SecA. Our results suggest a mechanism for nascent substrate protein recognition.
Data availability
X-ray crystallography data are deposited in PDB under accession code 6GOX.Small-angle x-ray scattering data are deposited in SASBDB under accession codes SASDDY9, SASDDZ9 and SASDE22.
Article and author information
Author details
Funding
Biotechnology and Biological Sciences Research Council (BB/L019434/1)
- Mohammed Jamshad
- Damon Huber
Biotechnology and Biological Sciences Research Council (BB/P009840/1)
- Timothy J Knowles
- Gareth W Hughes
Biotechnology and Biological Sciences Research Council (MIBTP)
- Max Wynne
Deutsche Forschungsgemeinschaft (FOR 1805)
- Günter Kramer
- Bernd Bukau
Deutsche Forschungsgemeinschaft (SFB 638)
- Günter Kramer
- Bernd Bukau
Wellcome (099266/Z/12/Z)
- Fiyaz Mohammed
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2019, Jamshad et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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The C-terminal tail of the bacterial translocation ATPase SecA modulates its activity
eLife 8:e48385.
https://doi.org/10.7554/eLife.48385
Further reading
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- Biochemistry and Chemical Biology
- Microbiology and Infectious Disease
Teichoic acids (TA) are linear phospho-saccharidic polymers and important constituents of the cell envelope of Gram-positive bacteria, either bound to the peptidoglycan as wall teichoic acids (WTA) or to the membrane as lipoteichoic acids (LTA). The composition of TA varies greatly but the presence of both WTA and LTA is highly conserved, hinting at an underlying fundamental function that is distinct from their specific roles in diverse organisms. We report the observation of a periplasmic space in Streptococcus pneumoniae by cryo-electron microscopy of vitreous sections. The thickness and appearance of this region change upon deletion of genes involved in the attachment of TA, supporting their role in the maintenance of a periplasmic space in Gram-positive bacteria as a possible universal function. Consequences of these mutations were further examined by super-resolved microscopy, following metabolic labeling and fluorophore coupling by click chemistry. This novel labeling method also enabled in-gel analysis of cell fractions. With this approach, we were able to titrate the actual amount of TA per cell and to determine the ratio of WTA to LTA. In addition, we followed the change of TA length during growth phases, and discovered that a mutant devoid of LTA accumulates the membrane-bound polymerized TA precursor.
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