Sato M, Sasaki T, Kobayashi M, Kise H
Institute of Materials Science, University of Tsukuba, Ibaraki, Japan.
Biosci Biotechnol Biochem. 2000 Dec;64(12):2552-8. doi: 10.1271/bbb.64.2552.
Secondary structure of alpha-chymotrypsin in water/ethanol was investigated by circular dichroic (CD) spectroscopy. The changes in catalytic activity were discussed in terms of structural changes of the enzyme. Alpha-chymotrypsin formed beta-sheet structure in water/ethanol (50/50 by volume), but it was substantially less active as compared to that in water. At water/ethanol 10/90, alpha-chymotrypsin took on a native-like structure, which gradually changed to beta conformation with concomitant loss of activity. Change of solvent composition from water/ethanol 50/50 to 90/10 or 10/90 by dilution with water or ethanol, respectively, led to partial recovery of native or native-like structure and activity. In water/methanol, alpha-chymotrypsin tended to form stable beta-sheet structure at water/methanol ratios lower than 50/50, but the catalytic activity decreased with time. Change to alpha-helix structure with substantial loss in catalytic activity was observed when alpha-chymotrypsin was dissolved in water/2,2,2-trifluoroethanol with water contents lower than 50%. In water/2,2,2-trifluoroethanol 90/10, alpha-chymotrypsin initially had the CD spectrum of native structure, but it changed with time to that characteristic of beta-sheet structure.
通过圆二色光谱法研究了α-胰凝乳蛋白酶在水/乙醇中的二级结构。从酶的结构变化方面讨论了催化活性的变化。α-胰凝乳蛋白酶在水/乙醇（体积比50/50）中形成β-折叠结构，但与在水中相比，其活性显著降低。在水/乙醇10/90时，α-胰凝乳蛋白酶呈现出类似天然的结构，随着活性的丧失，该结构逐渐转变为β-构象。分别用水或乙醇稀释，将溶剂组成从水/乙醇50/50变为90/10或10/90，会导致天然或类似天然结构及活性的部分恢复。在水/甲醇中，当水/甲醇比例低于50/50时，α-胰凝乳蛋白酶倾向于形成稳定的β-折叠结构，但催化活性随时间降低。当α-胰凝乳蛋白酶溶解于水含量低于50%的水/2,2,2-三氟乙醇中时，观察到其转变为α-螺旋结构，同时催化活性大幅丧失。在水/2,2,2-三氟乙醇90/10中，α-胰凝乳蛋白酶最初具有天然结构的圆二色光谱，但随着时间推移，其转变为β-折叠结构的特征光谱。
