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31.
Comparison of activity and conformation changes during refolding of urea-denatured creatine kinase 总被引:5,自引:0,他引:5
The course of the recovery of the enzymatic activity and the native conformation during the renaturation of urea-denatured creatine kinase (ATP:creatine N-phosphotransferase, EC 2.7.3.2) has been studied. Under suitable conditions, an activity recovery of 95% can be obtained and the reactivation follows a triphasic course. The initial two phases are relatively fast, whereas the slow phase takes some 24 h to reach completion. The recovery of the native conformation has been followed by changes in fluorescence, ultraviolet absorption and in exposed SH groups and has been shown to be a biphasic process. Both the reactivation and the refolding processes are independent of protein concentrations within a certain range, showing that the dimerization of the enzyme molecule is not rate-limiting. A comparison of the rate constants for the refolding of the molecule with those for the recovery of its catalytic activity shows that these are not synchronized and the activity recovery approaches completion after the refolding and dimerization of the subunits so far as can be detected by the methods employed. The final stage of refolding with complete activity recovery probably involves subtle conformational changes of the dimeric enzyme molecule not detectable by the physiochemical methods used in the present study. 相似文献
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Mu-Chin Tzeng Ming Jhy Hseu Jun Hai Yang Richard John Guillory 《The protein journal》1986,5(3):221-228
Snake presynaptic toxins such as crotoxin, β-bungarotoxin and taipoxin block neuromuscular transmission through inhibiting the release of acetylcholine by their phospholipase A2 activities. On the other hand, many other phospholipase A2s show little neurotoxicity. It is likely that the difference lies in whether high affinity binding to nerve cell membranes exists or not. To test this idea, crotoxin, β-bungarotoxin and taipoxin were first radioactively labeled with Na(125I) without loss of their neurotoxicity. Using the radioactive toxins we have found that each of the three showed specific binding to synaptosomal membranes from guinea pig brain. In contrast, we could not detect specific binding of a non-neurotoxic pancreatic phospholipase A2. Crotoxin and taipoxin, but not β-bungarotoxin, also bound specifically to membrane preparation from other tissues. The binding of each toxin was not greatly affected by the other two toxins. The photoaffinity labeling technique has been used to obtain further information about the components which bind crotoxin. For this purpose, (125I) crotoxin was derivatized with N-hydroxysuccinimidyl-4-azidobenzoate. Autoradiographic analysis of the membranes following photoirradiation in the presence of the modified crotoxin revealed that an 85K dalton component was preferentially covalently conjugated with the crotoxin analogue in a specific manner. 相似文献
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氨基酰化酶中金属锌离子的功能作用 总被引:1,自引:0,他引:1
氨基酰化酶是含锌金属酶。该酶每摩尔蛋白中含2摩尔Zn(Ⅱ)离子。金属鳌合剂与酶作用,通过竞争螯合Zn(Ⅱ)离子使酶活力下降。残余活力与残留金属含量呈正相关。竞争螯合的结果,生成不含金属的脱辅基酶蛋白,并导致酶活力的丧失。脱辅基酶由于加入Zn(Ⅱ)离子而恢复其活力。实验表明金属锌离子是氨基酰化酶催化活力所必需。与Zn(Ⅱ)离子相似,Co(Ⅱ)离子也可与脱辅基酶相结合并使之复活。 在190—240nm区域内对比了天然酶、脱辅基酶蛋白与Co(Ⅱ)置换氨基酰化酶的圆二色谱。远紫外圆二色谱表明,与天然酶相比,在脱辅基酶中由于金属离子的丧失导致主链构象发生变化,其中α螺旋增加约7%。因而锌离子(钴离子)对蛋白主链的反应最适构象有一定的稳定作用。脱辅基酶与Co(Ⅱ)离子结合,酶的主链构象恢复至与天然酶几近相同。可认为这是促使酶复活的内在因素。 相似文献
36.
在两柄类胚胎初级诱导的研究中,诱导物质如何进入反应细胞是一个受到重视(Grunzand Staubach,1979;Toivonen,1979)、但是仍未能解决的问题。为了对这一问题有所阐明,进行了离体诱导实验和电镜的观察。东方蝾螈(Cynops orientalis)早期原肠胚的外胚层块用豚鼠骨髓粗提液(BME)——有效的中胚层诱导物(Toivonen,1953)——处 相似文献
37.
贵州产宽叶缬草(Valeriana officinalis L.var.latifolia Miq.)油,用Finnigan4510型毛细管气相色谱/质谱/计算机联用方法进行了化学成分分析,共检出了29个成分鉴定了其中21个成分,占全精油的92.33%,主要成分为乙酸龙脑酯,α-蒎烯,莰烯,β-蒎烯,柠檬烯,乙酸葛缕酯,二氢乙酸葛缕酯等。该油芬芳,适于调配烟用香精,亦用于调节器配食用香精和化妆品香精。 相似文献
38.
Comparative studies were performed on the activities of superoxide dismutases, catalase and glutathione peroxidase in organ homogenates from three omnivorous fishes, the barbel, crucian carp and common carp. The lipid peroxidation and protein contents of organ homogenates were also compared. These comparative measurements primarily provide control values for subsequent toxicological examinations. The highest total superoxide dismutase activities were found in the liver or roe, kidney, heart and spleen in every cases. The antioxidant enzyme activities and other studied parameters of the organ homogenates partly appear to depend on the feeding mode, but are rather characteristic of the fish variety. 相似文献
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