Transport of Small Molecules across the Bacterial Translocation Channel SecYEG

Abstract

Many proteins are translocated across the endoplasmic reticulum (ER) membrane or the bacterial plasma membrane through a conserved channel, formed by a heterotrimeric protein complex (called the Sec61p complex in eukaryotes and the SecYEG complex in bacteria and archaea). Cotranslational or postranslational translocation requires ribosome or SecA binding, respectively. The resting channel is impermeable to ions and water (Saparov et al., 2007). Here we have tested whether the channel remains a barrier to small molecules after a conformational transition occurred due to ligand binding. Therefore, we reconstituted the purified and fluorescently labeled translocation channel SecYEG into planar lipid bilayers. Positioning of the membrane on top of a laser scanning microscope with single molecule sensitivity allowed monitoring the reconstitution efficiency. Both the motor molecule SecA and ribosomes induced ion channel activity. The probability of the channel opening was derived from the number of open channels and the total number of reconstituted channels.Acknowledgments: The project was funded by the Austrian Science Fund.ReferenceS. M. Saparov, Karl Erlandson, Kurt Cannon, Julia Schaletzky, Sol Schulman, Tom A. Rapoport, and P. Pohl. Determining the Conductance of the SecY Protein Translocation Channel for Small Molecules. Mol.Cell 26 (4):501-509, 2007.