Transport and polymerisation of bacterial polysaccharides: from cytoplasm to the outside world. (360G-Wellcome-100209_Z_12_Z)

Extracellular polysaccharides play a variety of roles in bacteria, most relevant to the Trust is their role in the pathogenesis of bacteria. The sugar polymers can help evade the immune system, protect against the immune response or even modulate the immune system. I wish to study the process by which sugar molecules are transported across the cytoplasmic membrane, polymerized and attached to the proteins. There are of course conceptual similarities to protein glycosylation in eukaryotes. The fi rst step of the process is coupling of sugar to a lipid carrier. There are two broad classes of integral membrane proteins that carry out this process, one of which has a similarity to human proteins. We have expressed both these integral membrane protein and propose to study both its structure and mechanism. The next step is flipping across the cytoplasm, carried out by the flippase protein. From a biochemical viewpoint this is a fascinating reaction and distinct from the ATP driven process of MsbA etc. The units are then polymerized into chain of a defined length by the polymerase, another integral membrane protein. The length of the polymer is tightly regulated and we have data to suggest a possible molecular mechanism for this. The polymer can be attached to a protein or exported or transferred to another receptor. Our focus will be on the attachment of the polymer to protein substrates. We have cloned and expressed two such transferases and have crystals of one which diffract to a round 5A.