Mesures des déclins de l'anisotropie de fluorescence de la γ‐globuline et de ses fragments Fab, Fc et F(ab)2 marqués avec le l‐sulfonyl‐5‐diméthyl‐aminonaphtalène
Mesures des déclins de l'anisotropie de fluorescence de la γ‐globuline et de ses fragments Fab, Fc et F(ab)2 marqués avec le l‐sulfonyl‐5‐diméthyl‐aminonaphtalène
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Fab、Fc 和 F(ab)2 分别为 L-磺酰基-5-二甲基-氨基萘
DOI：
10.1111/j.1432-1033.1972.tb01662.x
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发表时间：
1972
期刊：
影响因子：
5.4
通讯作者：
P. Wahl
中科院分区：
文献类型：
--
作者：
J. Brochon;P. Wahl
用很短的闪光激发荧光的两个偏振分量的时间历程，测定了丹酚化γ球蛋白、Ig G及其酶片段[Fab，Fc，F(Ab)2]的总荧光和荧光各向异性衰减。
总荧光的衰变可以用三个指数之和来解释。各向异性衰减曲线可以用两个弛豫过程来解释。第一个过程以弛豫θC为特征，非常短，对应于小分子碎片的局部运动。第二个过程，弛豫时间θL，是长的，是由于分子的全局运动。在衰变曲线的末尾，θL的影响占主导地位。
关于FAB和Fc碎片的这部分曲线可以用等效椭球的模型来解释，其大小与在低角X射线散射测量中发现的相似。
对Ig G和F(Ab)2的各向异性衰变的分析表明，F(Ab)2和Fc片段的结合部分没有很大的旋转自由度。我们的结果不允许我们证实分子的这一部分存在一些有限的旋转自由，正如Yguerabere，Epstein和Stryer[10]所提出的那样。
最后，我们讨论了经典的荧光退偏方法在使用连续激发时的意义。结果表明，在T/θ的整个实验范围内，退偏曲线的斜率受ηC变化的影响。因此，用这种方法不可能直接得到免疫球蛋白及其片段的大分子弛豫时间θL。
The time course of the two polarized components of the fluorescence excited by a very short flash has been used to determine the total fluorescence and the fluorescence anisotropy decays of dansylated γ-globulin, IgG and its enzymic fragments [Fab, Fc, F(ab)2].
The decays of the total fluorescence can be accounted for by the sums of three exponentials. The anisotropy decay curves can be interpreted by two relaxation processes. The first process, characterized by the relaxation θC, is very short and corresponds to a local motion of small fragments of the molecules. The second process, of relaxation time θL, is long and is due to a global motion of the molecules. The influence of θL is predominant in the final part of the decay curves.
This part of the curve concerning Fab and Fc fragments can be interpreted by the model of the equivalent ellipsoid of about a similar size as that found in low angle X-ray scattering measurements.
The analysis of the anisotropy decays of IgG and of F(ab)2 do not reveal a great freedom of rotation in the joint portion of the F(ab)2 and Fc fragments. Our results do not allow us to confirm the presence of some limited freedom of rotation in that part of the molecule, as has been proposed by Yguerabide, Epstein and Stryer [10].
Finally, we discuss the significance of the classical method of fluorescence depolarization where one uses a continuous excitation. It is shown that the slope of the depolarization curve is influenced by the variation of θC in the whole experimental range of T/η. Consequently, it is not possible to obtain directly the macromolecular relaxation time θL of the molecules of IgG and its fragments, using this method.