Impact of fluorescent protein fusions on the bacterial flagellar motor

Impact of fluorescent protein fusions on the bacterial flagellar motor

Heo, M. (Centre de Biochimie Structurale)
Nord, A. L. (Centre de Biochimie Structurale)
Chamousset, D. (Centre de Biochimie Structurale)
van Rijn, E. (TU Delft BN/Technici en Analisten)
Beaumont, H.J.E. (TU Delft BN/Bertus Beaumont Lab)
Pedaci, F. (Centre de Biochimie Structurale)

2017

Fluorescent fusion proteins open a direct and unique window onto protein function. However, they also introduce the risk of perturbation of the function of the native protein. Successful applications of fluorescent fusions therefore rely on a careful assessment and minimization of the side effects, but such insight is still lacking for many applications. This is particularly relevant in the study of the internal dynamics of motor proteins, where both the chemical and mechanical reaction coordinates can be affected. Fluorescent proteins fused to the stator of the Bacterial Flagellar Motor (BFM) have previously been used to unveil the motor subunit dynamics. Here we report the effects on single motors of three fluorescent proteins fused to the stators, all of which altered BFM behavior. The torque generated by individual stators was reduced while their stoichiometry remained unaffected. MotB fusions decreased the switching frequency and induced a novel bias-dependent asymmetry in the speed in the two directions. These effects could be mitigated by inserting a linker at the fusion point. These findings provide a quantitative account of the effects of fluorescent fusions to the stator on BFM dynamics and their alleviation-new insights that advance the use of fluorescent fusions to probe the dynamics of protein complexes.

http://resolver.tudelft.nl/uuid:60a7431d-155b-42f9-9d41-31b51001e251

https://doi.org/10.1038/s41598-017-11241-w

2045-2322

Scientific Reports, 7 (1)

Institutional Repository

journal article

© 2017 M. Heo, A. L. Nord, D. Chamousset, E. van Rijn, H.J.E. Beaumont, F. Pedaci