Molecular Mechanisms of Marine Polysaccharide Depolymerization and Metabolism.

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Ocean plant biomass [i.e. microalgae and macroalgae (seaweed)] is estimated to be ~1/200 that of terrestrial plant biomass yet the mass-normalized annual turnover rate of photosynthetically fixed carbon in the oceans is roughly two-orders of magnitude faster than on land. This extremely dynamic environment is therefore very important to the global carbon cycle. The cell wall and storage polymers of seaweed, which are the bulk of seaweed biomass, comprise a group of complex polysaccharides that are structurally different from those of terrestrial plants. This photosynthetically fixed carbon is returned to the global carbon cycle primarily through the metabolic action of microbes but the microbiology and biochemistry underlying the process remains poorly studied. **The overarching objective of this research program is to address how microbes depolymerize seaweed polysaccharides with the ultimate aim of providing molecular level detail for the pathways required to metabolize seaweed polysaccharides. The specific objectives are to:*1. Isolate uncharacterized seaweed polysaccharide degrading bacteria, sequence their genomes, then exploit these genomes with existing genomes to delineate potential seaweed polysaccharide metabolism pathways.*2. Analyse the specific and/or genome-wide expression of genes in selected bacteria in response to seaweed biomass or specific polysaccharides.*3. Use a structure-function approach to uncover the molecular details behind the enzymatic bioconversion of seaweed polysaccharides by bacteria.**Our initial approach will be to isolate aerobic seaweed degrading marine bacteria from local waters and exploit modern DNA sequencing technologies (e.g. Illumin HiSeq) to sequence their genomes. In addition to existing genomic, these new genomes will provide a rich resource to mine using bioinformatics approaches (e.g. automated genome annotation, annotation of carbohydrate-active enzymes with dbCAN) to give a broad view of the enzyme systems required to process seaweed polysaccharides (Objective 1). The genomic information will then be exploited using methods to analyse gene expression (qPCR, RNASeq, or microarray analysis) in response to particular seaweed biomass as growth substrates, such as intact seaweed or purified polysaccharides (Objective 2). Finally, the largest and most intensive objective that is receiving the bulk of our focus is to use a combination of molecular biology, recombinant protein expression, protein biochemistry, enzymology, and structural biology (primarily X-ray crystallography) to fully understand the molecular details of seaweed polysaccharide depolymerisation and utilization of the components (Objective 3).**This research program will provide detailed insight into the fundamentals of how microbes degrade seaweed polysaccharides. This is critical to unravelling how photosynthetically fixed carbon is recycled in our oceans and incorporating biochemical information into biogeochemical models of the global carbon cycle. Additionally, in a world that is seeing rising levels of greenhouse gases and climate change, eyes are being increasingly turned to "green" science, renewable resources, carbon neutral footprints, and alternative fuel sources. It has been generally overlooked but seaweed is a renewable resource whose cell wall and storage polysaccharides represent an attractive carbohydrate source for the production of bio-fuels, with many extremely appealing advantages over terrestrial feedstocks. Ultimately, the proposed research program will generate a knowledge platform that can be leveraged to design strategies for the enzymatic conversion of seaweed cell-wall polysaccharides into fermentable sugars that can be converted into biofuels.