Ying Q L, Kemme M, Simon S R
Department of Pathology, State University of New York at Stony Brook 11794.
Biochemistry. 1994 May 10;33(18):5445-50. doi: 10.1021/bi00184a013.
Secretory leukoprotease inhibitor (SLPI) comprises two homologous domains: the C-terminal domain contains the reactive site, while the function of the N-terminal domain remains unknown. In order to elucidate the function of the N-terminal domain, we studied the kinetics of reactions of human leukocyte elastase with two recombinant forms of SLPI: the full-length inhibitor and the C-terminal domain alone. The reactions of elastase with the full-length inhibitor and the C-terminal domain share the same association rate constant, 2 x 10(6) M-1 s-1, but the complex formed with the C-terminal domain is less stable, with a dissociation rate constant of 8 x 10(-4) s-1, 5 times higher than that of the complex with the full-length inhibitor. The binding of the full-length inhibitor to elastase is greatly accelerated by polyanions. In the presence of submicromolar concentrations (1 microgram/mL) of heparin, the association rate constant is increased by more than 1 order of magnitude. The binding of the C-terminal domain alone to elastase shows much lower sensitivity to heparin; in the presence of 5 microM (25 micrograms/mL) heparin, association of the C-terminal domain with elastase reaches a maximum rate of 7 x 10(6) M-1 s-1, about 3 times higher than the rate in the absence of heparin. Similar differential effects of heparin have been observed on the reactions of alpha-chymotrypsin with the two recombinant forms of SLPI.2=
分泌型白细胞蛋白酶抑制剂（SLPI）由两个同源结构域组成：C末端结构域包含反应位点，而N末端结构域的功能尚不清楚。为了阐明N末端结构域的功能，我们研究了人白细胞弹性蛋白酶与两种重组形式的SLPI的反应动力学：全长抑制剂和单独的C末端结构域。弹性蛋白酶与全长抑制剂和C末端结构域的反应具有相同的缔合速率常数，即2×10⁶ M⁻¹ s⁻¹，但与C末端结构域形成的复合物稳定性较差，解离速率常数为8×10⁻⁴ s⁻¹，比与全长抑制剂形成的复合物高5倍。多阴离子极大地加速了全长抑制剂与弹性蛋白酶的结合。在亚微摩尔浓度（1微克/毫升）的肝素存在下，缔合速率常数增加了一个多数量级。单独的C末端结构域与弹性蛋白酶的结合对肝素的敏感性要低得多；在5微摩尔（25微克/毫升）肝素存在下，C末端结构域与弹性蛋白酶的缔合达到最大速率7×10⁶ M⁻¹ s⁻¹，约为无肝素时速率的3倍。在α-胰凝乳蛋白酶与两种重组形式的SLPI的反应中也观察到了肝素的类似差异效应。
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