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C-terminal eYFP fusion impairs Escherichia coli MinE function

Palanisamy, Navaneethan
; 
Öztürk, Mehmet Ali
; 
Di Ventura, Barbara
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University of Freiburg; University of Heidelberg
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2020-01-10
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Chester Medical School
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Abstract
The Escherichia coli Min system plays an important role in the proper placement of the septum ring (Z-ring) at mid-cell during cell division. MinE forms a pole-to-pole spatial oscillator together with the membrane-bound ATPase MinD, which results in MinD having a concentration gradient with maxima at the poles and minimum at mid-cell. MinC, the direct inhibitor of the Z-ring initiator protein FtsZ, forms a complex with MinD at the membrane, thus mirroring MinD polar gradients. Therefore, MinC-mediated FtsZ inhibition occurs away from mid-cell. The existence of the oscillations was revealed by performing time-lapse microscopy with fluorescently-labeled Min proteins. These fusion proteins have been since then widely used to study properties of the Min system. Here we show that, despite permitting oscillations to occur in a range of protein concentrations, the enhanced yellow fluorescent protein (eYFP) C-terminally fused to MinE impairs its function. Combining in vivo, in vitro and in silico approaches, we demonstrate that the eYFP compromises MinE ability to displace MinC from MinD, to stimulate MinD ATPase activity and to directly bind to the membrane. Moreover, we reveal that MinE-eYFP is prone to aggregation. Taken together, our results indicate that this fusion is functionally impaired and should be used with caution in cell biological studies.
Citation
Palanisamy, N., Öztürk, M. A. & Di Ventura, B. (2020). C-terminal eYFP fusion impairs Escherichia coli MinE function. bioRxiv. https://doi.org/10.1101/2020.01.08.899393
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OpenRxiv (bioRxiv)
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http://hdl.handle.net/10034/629496
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10.1101/2020.01.08.899393
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Preprint
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https://www.biorxiv.org/content/10.1101/2020.01.08.899393v1