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All living cells are dependent on functional protein transport to survive. The essential ATPase SecA facilitates protein transport through the SecYEG translocon in Escherichia coli (E. coli). Until recently, it was believed that SecA only translocates proteins in a post-translational manner. However, new data suggest that SecA may also target proteins in a co-translational way. This project aimed to further investigate the cellular role of SecA with in vivo single-particle tracking (SPT). In vivo SPT allows us to directly measure the dynamics of a molecule of interest in living cells. On the basis of changes in diffusion, the prospect was to identify cytosolic SecA, SecYEG-bound SecA by the membrane, and possible ribosome-bound SecA using SPT.

The project identified that, in E. coli, SecA is mainly distributed along the cytoplasmic membrane, with a fraction of only 12% of SecA molecules diffusing freely in the cytosol. It was further estimated that SecA is mainly distributed along the cytoplasmic membrane in E. coli. I saw no evidence of the potential co-translational role of SecA. One of the major challenges during the project was that almost 90% of the SecA molecules were distributed by the membrane and 2D tracking could not pick up any dynamic changes by the membrane. Possible solutions to this challenge are addressed to be able to further elucidate the cellular role of SecA in the future.