Institute for Molecular Modeling and Simulation, University of Natural Resources and Life Sciences, 1190 Vienna, Austria.
Biochemistry. 2012 Oct 30;51(43):8636-53. doi: 10.1021/bi300802a. Epub 2012 Oct 16.
Cathepsins are mammalian papain-like cysteine proteases that play an important role in numerous physiological and pathological processes. In the present study, various molecular dynamics (MD) simulations of pro- and mature human cathepsins L and O were performed. This study is the first to report MD simulations to complement the initial model structure of (pro-)cathepsin O through conformational sampling, thus offering insight into the maturation of procathepsin O, which to date has not been described experimentally. The overall fold of (pro-)cathepsin O appears very similar to that of (pro-)cathepsin L. The propeptide binding loop (PBL)-propeptide interface of both procathepsins is found to form a stable two-stranded β-sheet. Additional stabilization of the PBL-propeptide interface is provided by hydrophobic side chain contacts in procathepsin L, whereas this seems to be due to charge-dipole interactions in procathepsin O. Introduction of two mutations (L147P and G148P) into procathepsin O entails a significant loss of hydrogen bonding, disabling formation of the interfacial β-sheet. Simulations at different protonation states suggest that procathepsin L is more sensitive to a change in pH than procathepsin O. Potential differences between the maturation of procathepsin O and procathepsin L inferred from the MD simulations might be caused by (i) stronger PBL-propeptide interactions in procathepsin O due to salt-bridge formation across the interface, (ii) more limited entropic gain of the propeptide of procathepsin O upon release into the bulk solvent due to diverse conformational states sampled in the bound state, (iii) more pronounced entropic loss of the PBL in procathepsin O upon substrate binding caused by diverse conformational states sampled in the free, mature enzyme, and (iv) lower sensitivity of procathepsin O to pH change caused by the presence of fewer carboxylate groups at the PBL-propeptide interface.
组织蛋白酶是哺乳动物木瓜蛋白酶样半胱氨酸蛋白酶，在许多生理和病理过程中发挥重要作用。本研究对原态和成熟的人组织蛋白酶 L 和 O 进行了各种分子动力学 (MD) 模拟。这是首次报道通过构象采样对（原）组织蛋白酶 O 的初始模型结构进行 MD 模拟，从而深入了解原组织蛋白酶 O 的成熟过程，这在实验上迄今尚未被描述过。（原）组织蛋白酶 O 的整体折叠结构与（原）组织蛋白酶 L 非常相似。两个原酶的前肽结合环 (PBL)-前肽界面被发现形成稳定的双链 β-折叠。在组织蛋白酶 L 中，PBL-前肽界面的额外稳定性是由疏水性侧链接触提供的，而在组织蛋白酶 O 中，这种稳定性似乎是由于电荷偶极相互作用。将两个突变（L147P 和 G148P）引入组织蛋白酶 O 会导致氢键显著丧失，从而无法形成界面 β-折叠。在不同质子化状态下的模拟表明，组织蛋白酶 L 对 pH 变化的敏感性比组织蛋白酶 O 更高。从 MD 模拟推断出的组织蛋白酶 O 成熟与组织蛋白酶 L 成熟之间的潜在差异可能是由以下原因引起的：(i) 由于界面处形成盐桥，组织蛋白酶 O 中 PBL-前肽相互作用更强；(ii) 由于结合态中采样的构象状态不同，组织蛋白酶 O 的前肽在释放到本体溶剂中时获得的熵增益有限；(iii) 由于在自由、成熟酶中采样的构象状态不同，组织蛋白酶 O 中的 PBL 在结合底物时熵损失更明显；(iv) 由于 PBL-前肽界面上的羧酸盐基团较少，组织蛋白酶 O 对 pH 变化的敏感性降低。
