Imperial College London
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DrMichaelHohl

Faculty of Medicine, Department of Infectious Disease

Research Associate
 
 
 
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Contact

 

m.hohl Website

 
 
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Location

 

Sir Alexander Fleming BuildingSouth Kensington Campus

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Summary

 

Publications

Citation

BibTex format

@article{Hürlimann:2017:10.1111/febs.14065,
author = {Hürlimann, LM and Hohl, M and Seeger, MA},
doi = {10.1111/febs.14065},
journal = {FEBS J},
pages = {1672--1687},
title = {Split tasks of asymmetric nucleotide-binding sites in the heterodimeric ABC exporter EfrCD.},
url = {http://dx.doi.org/10.1111/febs.14065},
volume = {284},
year = {2017}
}
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RIS format (EndNote, RefMan)

TY  - JOUR
AB  - Many heterodimeric ATP-binding cassette (ABC) exporters evolved asymmetric ATP-binding sites containing a degenerate site incapable of ATP hydrolysis due to noncanonical substitutions in conserved sequence motifs. Recent studies revealed that nucleotide binding to the degenerate site stabilizes contacts between the nucleotide-binding domains (NBDs) of the inward-facing transporter and regulates ATP hydrolysis at the consensus site via allosteric coupling mediated by the D-loops. However, it is unclear whether nucleotide binding to the degenerate site is strictly required for substrate transport. In this study, we examined the functional consequences of a systematic set of mutations introduced at the degenerate and consensus site of the multidrug efflux pump EfrCD of Enterococcus faecalis. Mutating motifs which differ among the two ATP-binding sites (Walker B, switch loop, and ABC signature) or which are involved in interdomain communication (D-loop and Q-loop) led to asymmetric results in the functional assays and were better tolerated at the degenerate site. This highlights the importance of the degenerate site to allosterically regulate the events at the consensus site. Mutating invariant motifs involved in ATP binding and NBD closure (A-loop and Walker A) resulted in equally reduced transport activities, regardless at which ATP-binding site they were introduced. In contrast to previously investigated heterodimeric ABC exporters, mutation of the degenerate site Walker A lysine completely inactivated ATPase activity and substrate transport, indicating that ATP binding to the degenerate site is essential for EfrCD. This study provides novel insights into the split tasks of asymmetric ATP-binding sites of heterodimeric ABC exporters.
AU  - Hürlimann,LM
AU  - Hohl,M
AU  - Seeger,MA
DO  - 10.1111/febs.14065
EP  - 1687
PY  - 2017///
SP  - 1672
TI  - Split tasks of asymmetric nucleotide-binding sites in the heterodimeric ABC exporter EfrCD.
T2  - FEBS J
UR  - http://dx.doi.org/10.1111/febs.14065
UR  - https://www.ncbi.nlm.nih.gov/pubmed/28417533
VL  - 284
ER  -
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