Diffusion of insulin-like growth factor-I and ribonuclease through fibrin gels
Diffusion of insulin-like growth factor-I and ribonuclease through fibrin gels
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DOI：
10.1529/biophysj.106.102699
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发表时间：
2007-06-01
影响因子：
3.4
通讯作者：
Anderson, John L.
中科院分区：
文献类型：
--
作者：
Nauman, Jess V.;Campbell, Phil G.;Anderson, John L.
本文介绍了一种同时测定两种蛋白质扩散系数的荧光法，并报道了胰岛素样生长因子（IGF-I）和核糖核酸酶（RNase）在0.27%纤维蛋白水凝胶中的扩散系数。该方法是基于双色成像的蛋白质浓度场随时间的松弛和比较的结果与运输模型。凝胶被限制在一个薄的（200 μ m）毛细管和蛋白质标记的荧光染料。实验测定的RNA酶扩散系数（D = 1.21 × 10（-6）cm（2）/s）与稀凝胶和散装水溶液的文献值一致，因此表明凝胶和染料对扩散的影响可忽略不计。在不与纤维蛋白基质结合的情况下，IGF-I的实验扩散系数（D = 1.59 x 10（-6）cm（2）/s）与X射线晶体学已知的分子尺寸以及基于14种其他蛋白质的D与分子量之间的相关性一致。在这里开发的实验方法持有承诺，用于确定在各种条件下，例如，当分子吸附到凝胶或对流通过凝胶的流体运输的生物分子的分子运输特性。
A fluorescence-based method for simultaneously determining the diffusion coefficients of two proteins is described, and the diffusion coefficient of insulin-like growth factor (IGF-I) and ribonuclease (RNase) in a 0.27% fibrin hydrogel is reported. The method is based on two-color imaging of the relaxation of the protein concentration field with time and comparing the results with a transport model. The gel is confined in a thin (200 mu m) capillary and the protein is labeled with a fluorescent dye. The experimentally determined diffusion coefficient of RNase (D = 1.21 x 10(-6) cm(2)/s) agrees with literature values for dilute gels and bulk aqueous solutions, thus indicating the gel and the dye had a negligible effect on diffusion. The experimental diffusion coefficient of IGF-I (D = 1.59 x 10(-6) cm(2)/s), in the absence of binding to the fibrin matrix, is consistent with the dimensions of the molecule known from x-ray crystallography and a correlation between D and molecular weight based on 14 other proteins. The experimental method developed here holds promise for determining molecular transport properties of biomolecules under a variety of conditions, for example, when the molecule adsorbs to the gel or is convected through the gel by fluid transport.