Koo Bon-Hun, Park Michael Y, Jeon Ok-Hee, Kim Doo-Sik
National Research Laboratory, Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, 134 Sinchon-Dong Seodaemun-Gu, Seoul 120-749, Korea.
J Biol Chem. 2009 Aug 28;284(35):23375-85. doi: 10.1074/jbc.M109.036848. Epub 2009 Jul 6.
Matrix metalloprotease (MMP)-2 plays a key role in many biological and pathological processes related to cell migration, invasion, and mitogenesis. MMP-2 is synthesized as a zymogen that is activated through either a conformational change or proteolysis of the propeptide. Several activating enzymes for pro-MMP-2 have been proposed, including metalloproteases and serine proteases. The mechanism of pro-MMP-2 activation by metalloproteases is well established, and the most studied activation mechanism involves cleavage of the propeptide by membrane type 1-MMP (MT1-MMP). In contrast, serine protease activation has not been thoroughly studied, although studies suggest that MT1-MMP may be involved in activation by thrombin and plasmin. Here, we demonstrate that factor Xa mediates MT1-MMP-independent processing of pro-MMP-2 in vascular smooth muscle cells and endothelial cells. Factor Xa and thrombin directly cleaved the propeptide on the carboxyl terminal sides of the Arg(98) and Arg(101) residues, whereas plasmin only cleaved the propeptide downstream of Arg(101). Moreover, processed MMP-2 showed enzymatic activity that was enhanced by intermolecular autoproteolytic processing at the Asn(109)-Tyr peptide bond. In addition to its role in activation, factor Xa rapidly degraded MMP-2, thereby restricting excessive MMP-2 activity. Thrombin also degraded MMP-2, but the degradation was reduced greatly under cell-associated conditions, resulting in an increase in processed MMP-2. Overall, factor Xa and thrombin regulate MMP-2 enzymatic activity through its activation and degradation. Thus, the net enzymatic activity results from a balance between MMP-2 activation and degradation.
基质金属蛋白酶（MMP）-2在许多与细胞迁移、侵袭和有丝分裂相关的生物学和病理过程中起关键作用。MMP-2以酶原形式合成，通过前肽的构象变化或蛋白水解作用被激活。已提出几种前MMP-2的激活酶，包括金属蛋白酶和丝氨酸蛋白酶。金属蛋白酶激活前MMP-2的机制已得到充分证实，研究最多的激活机制涉及膜型1-MMP（MT1-MMP）对前肽的切割。相比之下，丝氨酸蛋白酶激活机制尚未得到深入研究，尽管研究表明MT1-MMP可能参与凝血酶和纤溶酶的激活过程。在此，我们证明因子Xa在血管平滑肌细胞和内皮细胞中介导不依赖MT1-MMP的前MMP-2加工过程。因子Xa和凝血酶直接在精氨酸（98）和精氨酸（101）残基的羧基末端侧切割前肽，而纤溶酶仅在精氨酸（101）下游切割前肽。此外，加工后的MMP-2表现出酶活性，该活性通过在天冬酰胺（109）-酪氨酸肽键处的分子间自蛋白水解加工而增强。除了其在激活中的作用外，因子Xa还能快速降解MMP-2，从而限制MMP-2的过度活性。凝血酶也能降解MMP-2，但在细胞相关条件下，降解作用大大降低，导致加工后的MMP-2增加。总体而言，因子Xa和凝血酶通过激活和降解来调节MMP-2的酶活性。因此，净酶活性是MMP-2激活和降解之间平衡的结果。
