Effects of ligand heterogeneity in the characterization of affinity columns by frontal analysis
Effects of ligand heterogeneity in the characterization of affinity columns by frontal analysis
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DOI：
10.1021/ac970565m
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发表时间：
1997-12-01
影响因子：
7.4
通讯作者：
Hage, DS
中科院分区：
文献类型：
--
作者：
Tweed, SA;Loun, B;Hage, DS
本文研究了在亲和柱中配体异质性对前沿分析法测定的结合容量和平衡常数的影响。推导的n-和双中心系统的方程与L-甲状腺素与人血清白蛋白（HSA）柱的结合以及（R）-华法林与HSA或鸽血清白蛋白偶联柱的结合结果吻合得很好。相同的方程被用来检查如何不同程度的配体异质性影响的表观结合能力或平衡常数使用双倒数前沿分析图的线性范围测量。很大一部分双站点系统给出了良好的估计（即，小于10-20%的误差）。所有三位点和四位点情况的较小但仍然可观的部分也产生了对这些值的良好估计。这项工作的结果不限于蛋白质亲和柱，但应适用于任何类型的固定相，具有明确的结合区域和相对快速，可逆的相互作用与溶质。
This study examined how the binding capacities and equilibrium constants measured by frontal analysis are affected by ligand heterogeneity in affinity columns, Equations derived for n- and two-site systems gave good agreement with results obtained for the binding of L-thyroxine to a column containing human serum albumin (HSA) and for the binding of (R)-warfarin to coupled columns containing HSA or pigeon serum albumin, The same equations were used to examine how different degrees of ligand heterogeneity affected the apparent binding capacities or equilibrium constants measured using the linear range of double-reciprocal frontal analysis plots. A large proportion of two-site systems gave good estimates (i.e., less than 10-20% error) for the true total column capacity and for the association constant of the highest affinity ligand in the column, A smaller, but still appreciable, fraction of all three-and four-site cases also produced good estimates of these values. The results of this work are not limited to protein-based affinity columns but should be applicable to any type of stationary phase that has well-defined binding regions and relatively fast, reversible interactions with solutes.