A fast protein liquid chromatography (FPLC) system for purification and characterization of DMSP metabolic enzymes

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This is a reapplication for funding to buy an ÄKTA Purifier fast protein liquid chromatography (FPLC) system that is urgently needed to replace a ~20 yr old obsolete, unsupported, and inoperable FPLC. FPLCs are an essential tool for purifying and characterizing enzymes (protein catalysts). My research program seeks to discover and characterize marine algal enzymes that produce the world's most abundant organosulfur compound, dimethylsulfoniopropionate (DMSP). DMSP is vital 1) for algal cellular protection; 2) to bacteria who prefer DMSP as a sulfur source; 3) for supplying half the sulfur for the global biogeochemical sulfur cycle; and 4) to climate regulation since DMSP-derived aerosols form clouds. I exploit 3 complementary strategies (enzyme purification, comparative genomics, bacterial functional complementation assays) to discover genes encoding DMSP metabolic enzymes.The latter two approaches identify DMSP gene candidates that we will express in bacterial hosts as recombinant enzymes which then require purification. Native algal DMSP enzymes will be purified to enable protein sequencing tools which will reveal the genes encoding DMSP enzymes. These strategies all *depend* on purified, high-quality enzymes in order to confirm and study their function. Purified DMSP enzymes will be characterized with respect to their identity, biochemical structure, regulation, and kinetic behaviour. DMSP gene identification will enable phylogenetic studies of how widespread DMSP production is in marine algae and can enable studies predicting the response of algal DMSP production to future oceanic conditions. The FPLC system will see substantial use by my group (at least 1-2 postdoctoral and 15 undergraduates over my current Discovery award) and other researchers at Mt. Allison University, providing excellent HQP training in advanced protein purification technologies and supporting gene discovery efforts.