Inactivation of chymotrypsin by 5-benzyl-6-chloro-2-pyrone: 13C NMR and X-ray diffraction analyses of the inactivator-enzyme complex.
Inactivation of chymotrypsin by 5-benzyl-6-chloro-2-pyrone: 13C NMR and X-ray diffraction analyses of the inactivator-enzyme complex.
复制标题
5-苄基-6-氯-2-吡喃酮对胰凝乳蛋白酶的灭活：灭活剂-酶复合物的 13C NMR 和 X 射线衍射分析。
DOI：
10.1021/bi00322a011
复制
发表时间：
1985
期刊：
影响因子：
2.9
通讯作者：
Abeles,RH
中科院分区：
文献类型：
--
作者：
Ringe,D;Seaton,BA;Gelb,MH;Abeles,RH
化学系，马萨诸塞州理工学院，剑桥，马萨诸塞州02139，和生物化学研究生部，布兰迪斯大学，沃尔瑟姆，马萨诸塞州02254接收于1984年3月13日;修订的Mandarin pt接收于1984年8月10日摘要：研究了5-苄基-6-氯-2-吡喃酮对胰凝乳蛋白酶的灭活作用。失活酶的氯化物分析表明，氯不再存在于复合物中。用5-苄基-6-氯-2-吡喃酮-2（d-13 C2）灭活的胰凝乳蛋白酶的13 C NMR光谱显示存在来自蛋白结合灭活剂的两个新共振。这些共振的化学位移值与酶上完整的吡喃酮环以及C-6氯被不同杂原子取代一致。X-射线衍射分析表明，在1.5倍分辨率的失活酶复合物的活性位点丝氨酸残基（丝氨酸-195）的-氧共价attachedto C-6的失活剂和吡喃酮环是完整的。灭活剂的5-苄基与疏水特异性口袋中的酶结合。本文讨论了由于与失活剂络合而引起的蛋白质构象变化，最近有报道称，6-氯-2-吡喃酮可使胰凝乳蛋白酶和其它丝氨酸蛋白酶失活（Westkaemper & Abeles，1983）。胰凝乳蛋白酶被5-苄基-6-氯-2-吡喃酮（1）和3-苄基-6-氯-2-吡喃酮（2）以时间依赖性方式灭活。
Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, and Graduate Department of Biochemistry, Brandéis University, Waltham, Massachusetts 02254 Received March 13, 1984; Revised Manuscript Received August 10, 1984 abstract: The inactivation of chymotrypsin by 5-benzyl-6-chloro-2-pyrone has been studied. Chloride analysis of the inactivated enzyme suggests that chlorine is no longer present in the complex. 13C NMR spectroscopy of chymotrypsin inactivated with 5-benzyl-6-chloro-2-pyrone-2, d-13C2 shows the presence of two new resonances from the protein-bound inactivator. The chemical shift values of these resonances are consistent with an intact pyrone ring on the enzyme as well as the replacement of the C-6 chlorine by a different heteroatom. X-ray diffraction analysis at 1.5-Á resolution of the inactivator-enzyme complex demonstrates that the-oxygen of the active site serine residue (serine-195) is covalently attachedto C-6 of the inactivator and that the pyrone ring is intact. The 5-benzyl group of the inactivator is bound to the enzyme in the hydrophobicspecificity pocket. The conformational changes that occur in the protein as a result of complexation with the inactivator are discussed.Xhe inactivation of chymotrypsin and other serine proteases by substituted 6-chloro-2-pyrones has recently been reported (Westkaemper & Abeles, 1983). Inactivation of chymotrypsin by 5-benzyl-6-chloro-2-pyrone (1) and 3-benzyl-6-chloro-2-pyrone (2) occurs in a time-dependent manner.