The twin arginine translocase (Tat) is a protein translocation system dedicated to the transport of folded proteins across the bacterial cytoplasmic membrane and the thylakoid membrane of plant chloroplasts. In E. coli, the integral membrane proteins TatA, TatB and TatC are essential components of the Tat pathway. Multiple copies of TatB and TatC are found in a TatBC complex that also contains small, but varying amounts, ofTatA. In previous work this complex has been shown to function as the substrate receptor of the Tat pathway, recognising and interacting with Tat substrates through their signal peptides. Purification studies were performed to isolate a homogenous preparation of TatBC suitable for structural characterisation. It was shown that purification in the absence of TatA did not affect the stability or size of the TatBC complex. By performing the membrane solubilisation and purification in the mild detergent digitonin, TatBC could be characterised by blue-native PAGE. This identified four distinct complexes, the most abundant of which had an apparent molecular mass of approximately 430 kDa This complex was analysed by negative stain electron microscopy and found to exist as a roughly hemispherical structure with a small cavity in the centre. Sufi is an E. coli Tat substrate that has been extensively used to investigate the Tat pathway. No structural information exists for Sufi and its precise role in the cell remains unknown. The structure of Sufi was solved by X-ray crystallography at a resolution of 1.9 A. Analysis of the showed that, unlike its close structural homologue CueD, it is unable to bind copper atoms. Sequence analysis performed in the context of the structure identified several conserved regions that may play a role in Sufi function. By overproducing TatBC in conjunction with Sufi it was possible to isolate a TatBCsubstrate complex. This represents the first purification of an intermediate on the Tat pathway. It was shown that TaBC could bind two copies of Sufi, indicating the presence of multiple signal peptide binding sites. Negative stain electron microscopy studies showed that substrate binding occurred on the periphery of the TatBC complex with the substrate binding sites located adjacent to one another. The TatBC complex was observed to be smaller in the presence of bound Sufi, suggesting that it may undergo a significant conformational change upon substrate binding. Attempts were made to isolate other intermediates on the Tat pathway by using variant Tat or substrate proteins that were known to abolish Tat transport. No . complexes could be detected raising the possibility that the Tat system has a mechanism for disassembling stalled translocation machinery. Finally it was shown that TatAC complexes could be isolated in the absence of TatB. These appeared to contain an excess ofTatA over TatC and, intriguingly were able to bind substrate.