Enzyme specificity under dynamic control: A normal mode analysis of α-lytic protease
Enzyme specificity under dynamic control: A normal mode analysis of α-lytic protease
复制标题
DOI：
10.1006/jmbi.1998.2445
复制
发表时间：
1999-02
影响因子：
5.6
通讯作者：
David W. Miller;D. Agard
中科院分区：
文献类型：
--
作者：
David W. Miller;D. Agard
摘要 我们使用α-裂解蛋白酶作为模型系统来探索酶的内部动力学与其底物特异性之间的关系。野生型酶对其主要特异性口袋中的小底物具有高度特异性，而 M190A 突变体具有更广泛的特异性，可有效催化大底物和小底物的裂解。已经计算了野生型和突变型酶的正常模式，以确定内部振动如何影响这些对比的特异性特征。我们发现，对于排列在特异性口袋壁上的原子，野生型正常模式具有更加对称的特征，壁同相振动，并且口袋的大小保持相对固定。这与 120 K 下构象亚态的 X 射线晶体学数据一致。相反，我们发现在突变体中，结合袋正常模式具有更多的反对称特征，壁异相振动，并且袋能够膨胀和收缩。这些结果表明分子的内部振动可能在确定底物结合和特异性方面发挥重要作用。即使通过 NMR 弛豫或晶体学 B 因子测量的整体振动幅度基本保持不变，蛋白质结构的微小变化也会对分子振动模式产生重大影响，从而对酶性质产生重大影响。
Abstract We have used α-lytic protease as a model system for exploring the relationship between the internal dynamics of an enzyme and its substrate specificity. The wild-type enzyme is highly specific for small substrates in its primary specificity pocket, while the M190A mutant has a much broader specificity, efficiently catalyzing cleavage of both large and small substrates. Normal modes have been calculated for both the wild-type and the mutant enzyme to determine how internal vibrations contribute to these contrasting specificity profiles. We find that for the atoms lining the walls of the specificity pocket, the wild-type normal modes have a more symmetric character, with the walls vibrating in phase, and the size of the pocket remaining relatively fixed. This is in agreement with X-ray crystallographic data on conformational substates trapped at 120 K. In contrast, we find that in the mutant, the binding pocket normal modes have a more antisymmetric character, with the walls vibrating out of phase, and the pocket able to expand and contract. These results suggest that the internal vibrations of a molecule may play an important role in determining substrate binding and specificity. A small change in protein structure can have a significant effect on the pattern of molecular vibrations, and thus on enzymatic properties, even if the overall amplitudes of the vibrations, as measured by NMR relaxation or crystallographic B-factors, remain largely unchanged.