Structural and functional characterization of a novel serine protease responsible for activation of hepatocyte growth factor

肝细胞生长因子（HGF）是一种有效的肝细胞有丝分裂原，可能在肝损伤后的再生中起重要作用。HGF首先作为非活性单链前体合成，然后通过内源性蛋白水解加工转化为活性异二聚体。最近，一种新的丝氨酸蛋白酶（HGF激活剂）从血清中纯化出来。在本研究中，我们对该蛋白酶的结构和功能进行了分析，得到了以下结果：对HGF激活因子cDNA的分子克隆序列分析表明，从血清中纯化的HGF激活因子通过蛋白水解裂解从655个氨基酸的前体c端区衍生而来，该前体包含多个与凝血因子xii同源的结构域。HGF激活剂作为无活性的酶原存在于血浆中。在带负电荷的物质存在下，凝血酶可裂解Arg^<407>和Ile^<408>之间的键，从而激活酶原。因此，HGF激活因子酶原可能在组织损伤的凝血途径中被激活。正常大鼠肝、肾、肺、脾中HGF均为无活性单链。肝毒素处理后肝脏中HGF的生成明显增加。增加的HGF有很大一部分转化为活性形式，而在其他组织中没有发生这种转化。因此，在体内，蛋白水解激活系统作为一种将HGF活性定位到损伤组织的机制发挥作用。在损伤肝脏中检测到激活单链HGF的酶活性，而在正常肝脏中则没有。这种活性被抗hgf激活剂抗体抑制。因此，HGF激活剂可能是调控损伤组织中HGF活性的关键酶。

Hepatocyte growth factor (HGF) is a potent mitogen for heptocytes and may play an important role in regeneration following hepatic injury. HGF is first synthesized as an inactive single chain precursor which is then converted to an active heterodimeric from by endoproteolytic processing. Recently, a novel serine protease responsible for this processing (HGF activator) has been purified from serum. In this study, we analyzed the structure and function of this protease and obtained the following results.1. Molecular cloning sequence analysis of a cDNA for HGF activator revealed that HGF activator purified from serum is derived from the C-terminal region of a precursor of 655 amino acids by proteolytic cleavage and that the precursor consists of multiple domains homologous to those observed in blood coagulation factor XII.2. HGF activator was present in plasma as an inactive zymogen. The zymogen was activated by the cleavage of the bond between Arg^<407> and Ile^<408> by thrombin in the presence of negatively charged substances. Thus, the HGF activator zymogen may be activated during blood coagulation pathway in response to tissue injury.3. HGF remained as an inactive single chain from in the liver, kidney, lung and spleen of normal rat. The production of HGF markedly increased in the liver after hepatotoxin treatment. A significant portion of the increased HGF was converted to the active form, whereas the conversion did not occur in other tissues. Thus, the proteolytic activation system functions in vivo as a mechanism for localizing HGF activities to injured tissues.4. An enzymatic activity that activates single chain HGF was detected in the injured liver but not in the normal liver. This activity was inhibited by an anti-HGF activator antibody. Thus, HGF activator is likely to be the key enzyme regulating the activities of HGF in the injured tissues.