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 encoded by a different exon, raising the possibility that HSL is a mosaic protein. The catalytic serine (Ser423), as shown by site-directed mutagenesis, is encoded by exon 6. The phosphorylation site for cAMP-mediated activity control and a second site, which is presumably phosphorylated by 5' AMP-activated kinase, are encoded by exon 8, and a putative lipid-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 site 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 structure 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 characterization indicates that it has the same properties as HSL purified from rat adipose tissue.
Department of Medical and Physiological Chemistry, Lund University, Sweden.
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Holm
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Belfrage
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Osterlund
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Davis
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Schotz
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Langin
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D005075:Biological Evolution; D002478:Cells, Cultured; D017124:Conserved Sequence; D004950:Esterases / Q000235:genetics; D005091:Exons; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007438:Introns; D008954:Models, Biological; D009036:Moths / Q000166:cytology; D011994:Recombinant Proteins / Q000378:metabolism; D013045:Species Specificity; D002787:Sterol Esterase / Q000235:genetics* / Q000378:metabolism*
D011994:Recombinant Proteins; D004950:Esterases; D002787:Sterol Esterase