Conformational lability in serine protease active sites: Structures of hepatocyte growth factor activator (HGFA) alone and with the inhibitory domain from HGFA inhibitor-1B
Conformational lability in serine protease active sites: Structures of hepatocyte growth factor activator (HGFA) alone and with the inhibitory domain from HGFA inhibitor-1B
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DOI：
10.1016/j.jmb.2004.12.048
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发表时间：
2005-03-11
影响因子：
5.6
通讯作者：
Eigenbrot, C
中科院分区：
文献类型：
--
作者：
Shia, S;Stamos, J;Eigenbrot, C
肝细胞生长因子激活剂（HGFA）是一种丝氨酸蛋白酶，可将肝细胞生长因子（HGF）转化为活性形式。当活化的HGF与其同源受体Met结合时，细胞信号导致细胞生长、分化和迁移，促进肝脏、肾脏和皮肤组织再生。在HGF/Met活性与肿瘤发生相关的情况下，干预HGF向活性形式的转化有可能提供治疗益处。为了帮助确定调节HGF/Met效应的方法，我们确定了HGF/Met蛋白酶结构域的分子结构。我们在2.7埃分辨率下确定的结构，没有假底物或抑制剂结合，其特征是酶活性位点的关键残基具有非常规的构象。为了发现这种明显的非酶活性排列是否会在存在强相互作用抑制剂的情况下持续存在，我们还在2.6埃分辨率下确定了HGFA与生理抑制剂肝细胞生长因子激活因子抑制剂1B （HAI-1B）的第一个Kunitz结构域（KD1）络合的x射线结构。在这个复合体中，我们观察到一个重排的底物结合裂缝，它与其他丝氨酸蛋白酶的裂缝非常相似，表明它具有极端的构象动力学。我们还表征了KD1对16种丝氨酸蛋白酶的抑制作用，发现先前报道的HAI-1B完整细胞外区域的酶特异性仅存在于KD1中。我们发现HGFA、基质酶、hepsin、血浆钾激肽和胰蛋白酶被有效抑制，并利用复杂的结构来合理化这些结果的结构基础。(C) 2005 Elsevier Ltd版权所有。
Hepatocyte growth factor activator (HGFA) is a serine protease that converts hepatocyte growth factor (HGF) into its active form. When activated HGF binds its cognate receptor Met, cellular signals lead to cell growth, differentiation, and migration, activities which promote tissue regeneration in liver, kidney and skin. Intervention in the conversion of HGF to its active form has the potential to provide therapeutic benefit where HGF/Met activity is associated with tumorigenesis. To help identify ways to moderate HGF/Met effects, we have determined the molecular structure of the protease domain of HGFA. The structure we determined, at 2.7 Angstrom resolution, with no pseudo-substrate or inhibitor bound is characterized by an unconventional conformation of key residues in the enzyme active site. In order to find whether this apparently nonenzymatically competent arrangement would persist in the presence of a strongly-interacting inhibitor, we also have determined, at 2.6 Angstrom resolution, the X-ray structure of HGFA complexed with the first Kunitz domain (KD1) from the physiological inhibitor hepatocyte growth factor activator inhibitor 1B (HAI-1B). In this complex we observe a rearranged substrate binding cleft that closely mirrors the cleft of other serine proteases, suggesting an extreme conformational dynamism. We also characterize the inhibition of 16 serine proteases by KD1, finding that the previously reported enzyme specificity of the intact extracellular region of HAI-1B resides in KD1 alone. We find that HGFA, matriptase, hepsin, plasma kallikrein and trypsin are potently inhibited, and use the complex structure to rationalize the structural basis of these results. (C) 2005 Elsevier Ltd. All rights reserved.