Ugwu F, Van Hoef B, Bini A, Collen D, Lijnen H R
Center for Molecular and Vascular Biology, University of Leuven, Belgium.
Biochemistry. 1998 May 19;37(20):7231-6. doi: 10.1021/bi9728708.
Matrix metalloproteinase-3 (MMP-3, or stromelysin-1) specifically hydrolyzes the Glu143-Leu144 peptide bond in 45-kDa single-chain urokinase-type plasminogen activator (scu-PA) and in its two-chain (tcu-PA) derivative, yielding a 17-kDa NH2-terminal domain comprising the u-PA receptor (u-PAR) binding site and a 32-kDa COOH-terminal moiety containing the serine proteinase domain of u-PA. The conversion is completely abolished in the presence of the MMP inhibitors EDTA or 1,10-phenanthroline. Biospecific interaction analysis indicates that binding of MMP-3 occurs through the 32-kDa fragment. The 32-kDa fragment derived from scu-PA (scu-PA-32k) has a specific activity of </=500 IU/mg, but it can be activated with plasmin to a two-chain derivative (tcu-PA-32k) with a specific activity of 79 000 IU/mg. tcu-PA and tcu-PA-32k moieties derived from scu-PA-32k by plasmin or from tcu-PA by MMP-3 have comparable amidolytic activities toward the chromogenic substrate S-2444 (kcat/Km of 110 and 160 mM-1 s-1, respectively) and similar plasminogen activating activities in a coupled chromogenic substrate assay. Specific binding of the 17-kDa NH2-terminal domain to THP-1 monocytoid cells is completely abolished by competition with scu-PA but is not affected by scu-PA-32k (residual binding of 88 +/- 9% (mean +/- SEM; n = 3) with 25-fold molar excess). Thus, MMP-3 removes a functional NH2-terminal u-PAR-binding domain from u-PA without affecting its enzymatic properties.
基质金属蛋白酶-3（MMP-3，或基质溶解素-1）特异性水解45 kDa单链尿激酶型纤溶酶原激活剂（scu-PA）及其双链（tcu-PA）衍生物中的Glu143-Leu144肽键，产生一个包含u-PA受体（u-PAR）结合位点的17 kDa NH2末端结构域和一个包含u-PA丝氨酸蛋白酶结构域的32 kDa COOH末端部分。在MMP抑制剂EDTA或1,10-菲咯啉存在下，这种转化完全被消除。生物特异性相互作用分析表明，MMP-3的结合通过32 kDa片段发生。源自scu-PA的32 kDa片段（scu-PA-32k）的比活性≤500 IU/mg，但它可以被纤溶酶激活为比活性为79 000 IU/mg的双链衍生物（tcu-PA-32k）。通过纤溶酶从scu-PA-32k衍生的tcu-PA和tcu-PA-32k部分或通过MMP-3从tcu-PA衍生的部分对发色底物S-2444具有相当的酰胺水解活性（分别为110和160 mM-1 s-1的kcat/Km），并且在偶联发色底物测定中具有相似的纤溶酶原激活活性。17 kDa NH2末端结构域与THP-1单核细胞样细胞的特异性结合通过与scu-PA竞争而完全消除，但不受scu-PA-32k的影响（在25倍摩尔过量时残留结合为88±9%（平均值±标准误；n = 3））。因此，MMP-3从u-PA中去除了一个功能性的NH2末端u-PAR结合结构域，而不影响其酶学性质。
