Parry M A, Jacob U, Huber R, Wisner A, Bon C, Bode W
Max-Planck Institute of Biochemistry Department for Structural Research Am Klopferspitz 18a, 82152, Martinsried, Germany
Structure. 1998 Sep 15;6(9):1195-206. doi: 10.1016/s0969-2126(98)00119-1.
Trimeresurus stejnejeri venom plasminogen activator (TSV-PA) is a snake venom serine proteinase that specifically activates plasminogen. Snake venom serine proteinases form a subfamily of trypsin-like proteinases that are characterised by a high substrate specificity and resistance to inhibition. Many of these venom enzymes specifically interfere with haemostatic mechanisms and display a long circulating half-life. For these reasons several of them have commercial applications and are potentially attractive pharmacological tools.
The crystal structure of TSV-PA has been determined to 2.5 A resolution and refined to an R factor of 17.8 (R free, 24.4). The enzyme, showing the overall polypeptide fold of trypsin-like serine proteinases, displays unique structural elements such as the presence of a phenylalanine at position 193, a C-terminal tail clamped via a disulphide bridge to the 99-loop, and a structurally conserved Asp97 residue. The presence of a cis proline at position 218 is in agreement with evolutionary relationships to glandular kallikrein.
We postulate that Phe 193 accounts for the high substrate specificity of TSV-PA and renders it incapable of forming a stable complex with bovine pancreatic trypsin inhibitor and other extended substrates and inhibitors. Mutational studies previously showed that Asp97 is crucial for the plasminogenolytic activity of TSV-PA, here we identify the conservation of Asp97 in both types of mammalian plasminogen activator - tissue-type (tPA) and urokinase-type (uPA). It seems likely that Asp97 of tPA and uPA will have a similar role in plasminogen recognition. The C-terminal extension of TSV-PA is conserved among snake venom serine proteinases, although its function is unknown. The three-dimensional structure presented here is the first of a snake venom serine proteinase and provides an excellent template for modelling other homologous family members.
竹叶青蛇毒纤溶酶原激活剂（TSV-PA）是一种能特异性激活纤溶酶原的蛇毒丝氨酸蛋白酶。蛇毒丝氨酸蛋白酶构成了类胰蛋白酶样蛋白酶的一个亚家族，其特点是具有高底物特异性和抗抑制性。这些毒液酶中的许多都能特异性干扰止血机制，并具有较长的循环半衰期。由于这些原因，它们中的几种具有商业应用价值，并且是潜在的有吸引力的药理学工具。
已确定TSV-PA的晶体结构分辨率为2.5 Å，并精修至R因子为17.8（自由R因子为24.4）。该酶呈现出类胰蛋白酶样丝氨酸蛋白酶的整体多肽折叠结构，具有独特的结构元件，如193位存在苯丙氨酸、通过二硫键与99环相连的C末端尾巴以及结构保守的Asp97残基。218位存在顺式脯氨酸与与腺激肽释放酶的进化关系一致。
我们推测苯丙氨酸193决定了TSV-PA的高底物特异性，使其无法与牛胰蛋白酶抑制剂及其他延伸底物和抑制剂形成稳定复合物。先前的突变研究表明Asp97对TSV-PA的纤溶酶原激活活性至关重要，在此我们发现两种类型的哺乳动物纤溶酶原激活剂——组织型（tPA）和尿激酶型（uPA）中Asp97均保守。tPA和uPA的Asp97在纤溶酶原识别中似乎可能具有类似作用。TSV-PA的C末端延伸在蛇毒丝氨酸蛋白酶中是保守的，尽管其功能尚不清楚。此处呈现的三维结构是蛇毒丝氨酸蛋白酶的首个结构，为模拟其他同源家族成员提供了出色的模板。
