Evaluation of guanidine-based multimodal anion exchangers for protein selectivity and orthogonality
Evaluation of guanidine-based multimodal anion exchangers for protein selectivity and orthogonality
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基于胍的多模式阴离子交换剂的蛋白质选择性和正交性评估
DOI：
10.1016/j.chroma.2021.462398
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发表时间：
2021
影响因子：
4.1
通讯作者：
Cramer, Steven M.
中科院分区：
文献类型：
--
作者：
Koley, Sushmita;Altern, Scott H.;Vats, Mayank;Han, Xuan;Jang, Dongyoun;Snyder, Mark A.;Belisle, Chris;Cramer, Steven M.
在本论文中，我们研究了一类新的胍基多峰阴离子交换树脂的层析行为。首先用线性梯度层析和模型酸性蛋白质文库在pH值为5、6和7的条件下评价了这些树脂的选择性和蛋白质回收率。在高配基密度下，单胍基树脂表现出显著的回收率问题，而双胍基树脂除了两个蛋白质外，其余所有蛋白质的回收率都很高。此外，与低密度单胍树脂相比，双胍树脂表现出更强的pH依赖性选择性。低密度单胍树脂和双胍树脂的盐洗脱范围分别为0.250~0.621µM和0.162~0.828µM氯化钠。然后建立了一个定量构效关系模型来预测这些蛋白质在不同pH下在胍类原型上的洗脱行为，总训练和测试得分分别为0.88和0.85。此外，将这些配体固定在自组装单分子膜(SAM)上，进行了分子动力学模拟，以表征它们的构象偏好，并深入了解它们的色谱行为的分子基础。最后，一个新开发的框架被用来评估双胍树脂的分离性以及它与多模阳离子交换剂Nuvia cPrime的正交性。这项评估是使用第二个模型蛋白质文库进行的，该文库包括酸性和碱性蛋白质。分析结果表明，与疏水性较弱的商品Nuvia aPrime 4A相比，双胍原型具有更高的对分离度(0.73)和对增强系数(0.42)，对分离度和增强因子分别为0.57和0.22。这种新的多模式阴离子交换配体的选择性和正交性增强，可能为生物处理应用提供潜在的机会。
In this paper, we examined the chromatographic behavior of a new class of guanidine-based multimodal anion exchange resins. The selectivities and protein recoveries on these resins were first evaluated using linear gradient chromatography with a model acidic protein library at pH 5, 6 and 7. While a single-guanidine based resin exhibited significant recovery issues at high ligand density, a bis-guanidine based resin showed high recoveries of all but two of the proteins evaluated in the study. In addition, the bis-guanidine resin showed a more pH dependent selectivity pattern as compared to the low density single-guanidine resin. The salt elution range for the low density single-guanidine and bis-guanidine resins was also observed to vary from 0.250 to 0.621 M and 0.162 to 0.828 M NaCl, respectively. A QSAR model was then developed to predict the elution behavior of these proteins on the guanidine prototypes at multiple pH with overall training and test scores of 0.88 and 0.85, respectively. In addition, molecular dynamics simulations were performed with these ligands immobilized on a self-assembled monolayer (SAM) to characterize their conformational preferences and to gain insight into the molecular basis of their chromatographic behavior. Finally, a recently developed framework was employed to evaluate the separability of the bis-guanidine resin as well as its orthogonality to the multimodal cation exchanger, Nuvia cPrime. This evaluation was carried out using a second model protein library which included both acidic and basic proteins. The results of this analysis indicated that the bis-guanidine prototype exhibited both higher pair separability (0.73) and pair enhancement (0.42) as compared to the less hydrophobic commercial Nuvia aPrime 4A with pair separability and enhancement factors of 0.57 and 0.22, respectively. The enhanced selectivity and orthogonality of this new multimodal anion exchange ligand may offer potential opportunities for bioprocessing applications.