活性中心Active site

构成活性中心的残基在一级序列上可能相距很远，经折叠后在空间上聚在一起。活性中心只占酶分子体积的一小部分，通常具有疏水环境，以提高静电相互作用和排除水分子。结合部位决定底物专一性，催化部位提供酸碱、亲核或金属离子催化。活性中心与过渡态的结合比与底物更紧密，这是酶降低活化能的核心原理（鲍林 1946 年提出）。

Residues of the active site may be far apart in sequence but are brought together by folding. The site occupies a small fraction of the enzyme and often has a hydrophobic environment that strengthens electrostatic interactions and excludes water. The binding site determines substrate specificity, and the catalytic site supplies acid–base, nucleophilic or metal-ion catalysis. The active site binds the transition state more tightly than the substrate, the central principle of how enzymes lower activation energy, proposed by Pauling in 1946.

胰凝乳蛋白酶等丝氨酸蛋白酶的活性中心含“催化三联体”Ser195、His57、Asp102：天冬氨酸使组氨酸定向，组氨酸夺取丝氨酸羟基的质子，使丝氨酸氧成为强亲核试剂进攻底物肽键羰基，形成四面体中间体；“氧负离子洞”中的主链 N–H 稳定其负电荷。有机磷农药和神经毒剂与这个丝氨酸共价结合，使乙酰胆碱酯酶失活。

Serine proteases such as chymotrypsin have a catalytic triad of Ser195, His57 and Asp102: aspartate orients histidine, which takes the proton from serine's hydroxyl so that the serine oxygen attacks the substrate's peptide carbonyl as a strong nucleophile, forming a tetrahedral intermediate whose negative charge is stabilised by backbone N–H groups in the oxyanion hole. Organophosphate pesticides and nerve agents bond covalently to the corresponding serine and inactivate acetylcholinesterase.

通过定点突变、化学修饰、X 射线晶体学和底物类似物抑制剂可以确定活性中心的组成。药物设计常以活性中心为靶点，设计与过渡态相似的抑制剂，如 HIV 蛋白酶抑制剂和新冠主蛋白酶抑制剂奈玛特韦。“过渡态类似物”还用于诱导产生催化抗体。

Site-directed mutagenesis, chemical modification, X-ray crystallography and substrate-analogue inhibitors reveal which residues form the active site. Drug design often targets it with inhibitors that mimic the transition state, such as HIV protease inhibitors and the SARS-CoV-2 main-protease inhibitor nirmatrelvir. Transition-state analogues have also been used to raise catalytic antibodies.