Protein organizations underpinning Wzm/Wzt dependent bacterial polysaccharide production

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Bacteria sheath themselves in a coat of molecules made predominantly from complex polysaccharides (literally many-sugar; cellulose and starch are familiar examples). These coats then are then the dominant surface with which bacteria interact with their environment. For pathogenic bacteria, this is a major aspect of the way the host "sees" them, making these polysaccharides potentially useful as vaccines. These coats are also targeted by bacteria-killing viruses; as a result, bacteria have strong incentives to evolve new coat variants to escape such attacks, and there are thousands of coat variants known. The machinery that builds these polysaccharides, while still quite complex and diverse, recycles many of the key components and approaches. We aim to study this machinery, using a variety of tools, including x-ray crystallography which will allow us to see these machines in atom-by-atom detail. This will allow us to develop a detailed understanding of how these components work, and then test these ideas by perturbing the proteins using genetic tools. Our immediate target is the bacterium Rhizobium tropici, a useful organism that fixes nitrogen for beans and maize. This organism builds a polysaccharide coat incorporating the unusual sugars fucose and deoxytalose. We aim to investigate how these sugar adding enzymes work. This organism also measures its coat sugars to a fairly exact pre-specified length; however, the machinery building the saccharide chains lacks the typical organization seen in other saccharide measuring systems. We plan to investigate the measurement mechanism in some detail. In addition, the machinery adds an acetyl group to a subset of the deoxytalose residues. Published data hints that it may be doing so in such a way that only the top portion of the sugar is modified, helping thwart viruses which then would not recognize the bottom half. We want to test this hypothesis, and understand how such zonal control is achieved. Long term, we hope by understanding different such polysaccharide synthesis machines with unique properties, sufficient understanding will emerge to allow detailed predictions of the saccharide coat structures of all bacteria.