Understanding the TatA channel of the twin-arginine protein translocase

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Some bacterial proteins operate on the outside of the cell, for example the toxins produced by bacterial pathogens. Since all proteins are made inside the bacterium the extracellular proteins must be moved out of the cell across the normally impermeable cell membrane. This task is carried out by machines termed protein transporters that are located in the cell membrane. One type of transporter moves unfolded proteins, threading them across the membrane like string through the eye of a needle. By contrast, a second type of transporter, which we term the Tat system, moves folded proteins across the membrane. The Tat protein transport system is also found in plant chloroplasts. One essential component of the Tat system is the protein TatA. Multiple copies of TatA assemble to form a large complex. We have used electron microscopy to visualize the TatA complex. TatA forms a doughnut-shaped structure. This suggests that TatA forms a channel across the cell membrane with proteins being transported through the central hole. A surprising feature of TatA is that individual TatA complexes have different sizes, apparently because they contain different numbers of TatA proteins. It is possible that this size variation allows TatA to pack tightly around the protein that is being transported and thus prevent the co-transport of other molecules. However, the TatA complex may be flexible for a number of other plausible reasons. In this project we want to increase our understanding of how the TatA protein works. In particular we want to learn the significance of the variation in the size of the Tat complexes. The electron microscopy images are too fuzzy to allow us to determine the position or shape of individual proteins within the TatA complex. The organization of the subunits in the complex will instead be probed using methods that determine which parts of the TatA molecule are in contact with other TatA molecules by chemically tying the two proteins together. These links will also be used to determine which parts of the TatA protein have to move relative to other parts of the protein to allow TatA to carry out its function. We will investigate whether the variability in the size of the TatA complexes is linked to tailoring the size of the hole to the substrate by asking whether the size of the ring varies when transporting proteins of differing sizes. We will also test whether individual TatA proteins enter and leave the TatA complex during transport since this is a prediction of the substrate-fitting explanation of the size variations. These and other experiments are expected to give a clearer picture of the operation of TatA. The Tat system is a possible drug target because it is required for bacterial pathogenesis but is not found in humans. It is also of biotechnological interest because it could be used to secrete useful protein products.