Protein Translocation at the Single Molecule Level

Abstract

All proteins are synthesized in the cytoplasm. However, for many of them the cytoplasm is not their final destination. Instead, they need to be translocated across or into the plasma membrane. In bacteria, this process is mediated by the membrane-embedded protein-conducting SecYEG channel. This channel can either associate with the ribosome (co-translational translocation) or with a cytosolic motor protein, the SecA ATPase (post-tranlational translocation), with each the ability to provide the driving force for the translocation process across the membrane.Even though the SecYEG system has been intensively studied, many aspects of protein translocation remain elusive. Here, we study translocation by single-molecule fluorescence imaging. We reconstitute the bacterial SecYEG into phospholipid bilayer nanodiscs and immobilize these on a functionalized glass-surface. By monitoring the interactions of the components of the Sec translocon at resolution of individual molecules we aim to provide a means for better understanding the journey of proteins into or across the membrane.