(446c) Towards Understanding the Structure and Function of Alcohol Acetyltransferase Lipid Droplet Localization in S. Cerevisiae

Across eukaryotes metabolic activity of neutral lipid synthesis is localized to intracellular lipid droplet. Through specific protein domains, essential enzymes in the biosynthesis of triacylglycerols are associated with the outer membrane of the endoplasmic reticulum (ER) and ultimately to lipid droplets that emanate from the ER. Within these pathways, there are clear links between protein structure and function in terms of both lipid droplet association and enzymatic activity. In Saccharomyces cerevisiae, alcohol acetyltransferase 1 (ATF1) is also localized to lipid droplets. This enzyme catalyzes the reaction of acetyl-CoA and short chain alcohols to produce the corresponding ethyl esters, including the industrially useful solvent ethyl acetate and the flavor compound isoamyl ester (banana-like flavor and aroma). Compared to the triacylglycerol synthesis pathways there is little known about the structure of ATF1. The crystal structure has not been solved and there is no acceptable template from which an accurate homology model can be predicted. Here, we compare the sequence structure of ATF1 to proteins that are known to localize on lipid droplets and identify the domains responsible for localization. Truncation and localization studies reveal that both N- and C-terminal domains (amino acids 1-24 and 502-525) are necessary, but not sufficient, for ATF1 lipid droplet targeting. Similar to perilipin A, a structural protein localized to lipid droplets of adipocytes, internal hydrophobic domains of ATF1 are necessary for targeting. At present, the biological impetus for ATF1 localization is unknown. In this work we use our insights into the structure of ATF1 to explore the hypothesis that localization enhances ester synthesis pathway activity. We explore this hypothesis through comparative studies of ATF1 and cytosolic ATFs from Solanum lycopersicum, and the yeasts, Pichia anomala and Kluyveromyces lactis.