Physiological proteomics of relevant enzymes and transporters in marine polysaccharide-degrading Bacteroidetes

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Subproject A1 aims to characterize the physiology and adaptation of marine Bacteroidetes that specialize in polysaccharide degradation. We will functionally analyze molecular mechanisms which determine the success of key bacterial clades during phytoplankton blooms and their substrate specificity in polysaccharide utilization. The respective enzymes are encoded in dedicated genomic regions, so-called polysaccharide utilization loci (PULs). We use comparative quantitative subproteomic analyses of cultivable key bacteria under defined substrate conditions to deduce specific functions of selected PULs and their encoded CAZymes and transporters. In the focus of this project are bacterial isolates, which have been identified as relevant during spring phytoplankton blooms in the North Sea. The correlation of results obtained from defined laboratory cultivations on the one side and from environmental data on the other side is a crucial prerequisite to define ecological niches of key bacteria and their interaction in marine polysaccharide turnover in nature.In the first and second funding phase, subproject A1 focused on alpha- and beta-glucan specific degradation processes and determined key bacteria, PUL types and protein functions responsible for the utilization of these abundant marine sugars. Furthermore, we supported the detailed functional analyses of ulvan-, fucoidan-, xylan- and mannan- degradation processes in A2, A3, A4 and B1. Together with the consortium partners, we were able to determine major degradation pathways as well as new CAZyme functions of marine Bacteroidetes strains for these algal polysaccharides. During the third POMPU phase, we aim to finalize our detailed investigation of the cellular localization and putative functional interlinkages of laminarin-specific proteins in marine Bacteroidetes. We hypothesize that a strictly ordered protein machinery of sugar-binding, -degrading and -transporting proteins is a crucial molecular adaptation, which facilitates polysaccharide degradation in the highly diffusible marine environment. Since laminarin is the most abundant polysaccharide in marine habitats, we will continue to functionally analyze different types of laminarin-responsive PULs detectable in our Helgoland isolates and environmental samples during diatom-driven blooms. In this concluding POMPU phase, we will complete our detailed and systematic analyses of potential regulatory mechanisms, which determine a hierarchical expression of substrate-specific PULs and thus a controlled degradation of natural marine polysaccharides during phytoplankton blooms. We will furthermore finalize our functional and structural analyses of a new regulator type of laminarin PULs (discovered in the second POMPU phase), thus completing our investigation of these important marine PUL functions.