Lampidonis Antonis D, Argyrokastritis Alexandros, Stravopodis Dimitrios J, Voutsinas Gerassimos E, Ntouroupi Triantafyllia G, Margaritis Lukas H, Bizelis Iosif, Rogdakis Emmanuel
Department of Animal Science, Agricultural University of Athens, Iera Odos 75, 118 55 Athens, Greece.
Gene. 2008 Jun 15;416(1-2):30-43. doi: 10.1016/j.gene.2008.02.026. Epub 2008 Mar 12.
Hormone Sensitive Lipase (HSL) is a highly regulated enzyme that mediates lipolysis in adipocytes. HSL enzymatic activity is increased by adrenergic agonists, such as catecholamines and glucagons, which induce cyclic AMP (cAMP) intracellular production, subsequently followed by the activation of Protein Kinase A (PKA) and its downstream signalling cascade reactions. Since HSL constitutes the key enzyme in the regulation of lipid stores and the only enzyme being subjected to hormonal regulation [in terms of the recently identified Adipose Triglyceride Lipase (ATGL)], the ovine Hormone Sensitive Lipase (ovHSL) full-length cDNA clones were isolated, using a Polymerase Chain Reaction-based (PCR) strategy. The two isolated isoforms ovHSL-A and ovHSL-B contain two highly homologous Open Reading Frame (ORF) regions of 2.089 Kb and 2.086 Kb, respectively, the latter having been missed the 688th triplet coding for glutamine (DeltaQ(688)). The putative 695 and 694 amino acid respective sequences bear strong homologies with other HSL protein family members. Southern blotting analysis revealed that HSL is represented as a single copy gene in the ovine genome, while Reverse Transcription-PCR (RT-PCR) approaches unambiguously dictated its variable transcriptional expression profile in the different tissues examined. Interestingly, as undoubtedly corroborated by both RT-PCR and Western blotting analysis, ovHSL gene expression is notably enhanced in the adipose tissue during the fasting period, when lipolysis is highly increased in ruminant species. Based on the crystal structure of an Archaeoglobus fulgidus enzyme, a three-dimensional (3D) molecular model of the ovHSL putative catalytic domain was constructed, thus providing an inchoative insight into understanding the enzymatic activity and functional regulation mechanisms of the ruminant HSL gene product(s).
激素敏感脂肪酶（HSL）是一种受高度调控的酶，它介导脂肪细胞中的脂肪分解。肾上腺素能激动剂，如儿茶酚胺和胰高血糖素，可增加HSL的酶活性，这些激动剂诱导细胞内环状AMP（cAMP）的产生，随后激活蛋白激酶A（PKA）及其下游信号级联反应。由于HSL是调节脂质储存的关键酶，并且是唯一受激素调节的酶[就最近鉴定出的脂肪甘油三酯脂肪酶（ATGL）而言]，因此采用基于聚合酶链反应（PCR）的策略分离了绵羊激素敏感脂肪酶（ovHSL）全长cDNA克隆。分离出的两种同工型ovHSL-A和ovHSL-B分别包含两个高度同源的开放阅读框（ORF）区域，长度分别为2.089 kb和2.086 kb，后者缺失了编码谷氨酰胺的第688个三联体（DeltaQ(688)）。推测的695和694个氨基酸序列与其他HSL蛋白家族成员具有很强的同源性。Southern印迹分析表明，HSL在绵羊基因组中以单拷贝基因形式存在，而逆转录PCR（RT-PCR）方法明确显示其在不同检测组织中的转录表达谱存在差异。有趣的是，正如RT-PCR和蛋白质印迹分析所明确证实的那样，在反刍动物禁食期间脂肪组织中的脂肪分解高度增加时，ovHSL基因表达显著增强。基于嗜热栖热菌酶的晶体结构，构建了ovHSL推定催化结构域的三维（3D）分子模型，从而为理解反刍动物HSL基因产物的酶活性和功能调节机制提供了初步见解。
