HORMONE-SENSITIVE LIPASE - STRUCTURE, FUNCTION, EVOLUTION AND OVERPRODUCTION IN INSECT CELLS USING THE BACULOVIRUS EXPRESSION SYSTEM

Hormone-sensitive lipase (HSL) catalyses the rate-limiting step in the hydrolysis of stored triacylglycerols and is thereby a key enzyme in lipid metabolism and overall energy homeostasis. The gene organization of human HSL indicates that each putative functional region is encode d by a different exon, raising the possibility that HSL is a mosaic pr otein. The catalytic serine (Ser423), as shown by site-directed mutage nesis, is encoded by exon 6. The phosphorylation site for cAMP-mediate d activity control and a second site, which is presumably phosphorylat ed by 5' AMP-activated kinase, are encoded by exon 8, and a putative l ipid-binding region is encoded by the ninth and last exon. Besides the catalytic site serine motif (GXSXG), found in virtually all lipases, a sequence similarity between the region surrounding the catalytic sit e of HSL and that of five prokaryotic enzymes has been found, but the functional basis of this is not yet understood. To resolve the 3-D str ucture of HSL, an expression system utilizing recombinant baculovirus and insect cells has been established. The expressed protein, 80 mg/l culture, has been purified to homogeneity and a partial characterizati on indicates that it has the same properties as HSL purified from rat adipose tissue.