Abstract：
Snake venoms are known to contain toxins capable of interfering with normal physiological processes of victims. Specificity of toxins from snake venoms give scope to identify new molecules with therapeutic action and/or help to understand different cellular mechanisms. Russell\'s viper venom (RVV) is a mixture of many bioactive molecules with enzymatic and non-enzymatic proteins. The present article describes Daboialipase (DLP), an enzymatic phospholipase A2 with molecular mass of 14.3 kDa isolated from RVV. DLP was obtained after cation exchange chromatography followed by size-exclusion high performance liquid chromatography (SE-HPLC). The isolated DLP presented strong inhibition of adenosine di-phosphate (ADP) and collagen induced platelet aggregation. It also showed anti-thrombin properties by significantly extending thrombin time in human blood samples. Trypan blue and resazurin cell viability assays confirmed time-dependent cytotoxic and cytostatic activities of DLP on MCF7 breast cancer cells, in vitro. DLP caused morphological changes and nuclear damage in MCF7 cells. However, DLP did not cause cytotoxic effects on non-cancer HaCaT cells. Peptide sequences of DLP obtained by O-HRLCMS analysis showed similarity with a previously reported PLA2 (Uniprot ID: PA2B_DABRR/PDB ID: 1VIP_A). An active Asp at 49th position, calcium ion binding site and anticoagulant activity sites were identified in 1 VIP_A. These findings are expected to contribute to designing new anti-platelet, anticoagulant and anti-cancer molecules.
摘要：
已知蛇毒含有能够干扰受害者正常生理过程的毒素。来自蛇毒的毒素的特异性为识别具有治疗作用的新分子和/或帮助理解不同的细胞机制提供了空间。罗素的毒蛇毒液（RVV）是许多生物活性分子与酶和非酶蛋白的混合物。本文介绍了Daboialipase(DLP)，从RVV中分离的分子量为14.3kDa的酶促磷脂酶A2。在阳离子交换色谱、随后的尺寸排阻高效液相色谱(SE-HPLC)之后获得DLP。分离的DLP对二磷酸腺苷（ADP）和胶原蛋白诱导的血小板聚集具有很强的抑制作用。它还通过显著延长人血液样品中的凝血酶时间而显示抗凝血酶性质。台盼蓝和刃天青细胞活力测定证实了DLP对MCF7乳腺癌细胞的时间依赖性细胞毒性和细胞抑制活性，在体外。DLP引起MCF7细胞的形态学改变和核损伤。然而,DLP对非癌症HaCaT细胞没有引起细胞毒性作用。通过O-HRLCMS分析获得的DLP的肽序列显示与先前报道的PLA2(UniprotID:PA2B_DABRR/PDBID:1VIP_A)的相似性。第49位有一个活跃的Asp，在1个VIP_A中确定了钙离子结合位点和抗凝血活性位点。这些发现有望有助于设计新的抗血小板，抗凝血和抗癌分子。
