Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), 20 Biopolis Way, Centros #06-01, Singapore 138668.
Biotechnol Bioeng. 2012 Mar;109(3):736-46. doi: 10.1002/bit.24343. Epub 2011 Dec 28.
The use of linear PEGs for protein precipitation raises the issues of high viscosity and limited selectivity. This paper explores PEG branching as a way to alleviate the first problem, by using 3-arm star as the model branched structure. 3-arm star PEGs of 4,000 to 9,000 Da were synthesized and characterized. The effects of PEG branching were then elucidated by comparing the branched PEG precipitants to linear versions of equivalent molecular weights, in terms of IgG recovery from CHO cell culture supernatant, precipitation selectivity, solubility of different purified proteins, and precipitation kinetics. Two distinct effects were observed: PEG branching reduced dynamic viscosity; secondly, the branched PEGs precipitated less proteins and did so more slowly. Precipitation selectivity was largely unaffected. When the branched PEGs were used at concentrations higher than their linear counterparts to give similar precipitation yields, the dynamic viscosity of the branched PEGs were noticeably lower. Interestingly, the precipitation outcome was found to be a strong function of PEG hydrodynamic radius, regardless of PEG shape and molecular weight. These observations are consistent with steric mechanisms such as volume exclusion and attractive depletion.
线性聚乙二醇（PEG）用于蛋白质沉淀会带来高黏度和选择性有限的问题。本文探讨了 PEG 支化作为缓解第一个问题的方法，使用 3 臂星形作为模型支化结构。合成并表征了 4000 至 9000 Da 的 3 臂星形 PEG。然后，通过比较从 CHO 细胞培养上清液中回收 IgG、沉淀选择性、不同纯化蛋白的溶解度和沉淀动力学，研究了支化 PEG 沉淀剂相对于等效分子量的线性版本的支化效果。观察到两种明显的效果：PEG 支化降低了动态黏度；其次，支化 PEG 沉淀的蛋白较少，且沉淀速度较慢。沉淀选择性基本不受影响。当支化 PEG 的浓度高于线性 PEG 以获得相似的沉淀产率时，支化 PEG 的动态黏度明显降低。有趣的是，无论 PEG 形状和分子量如何，沉淀结果均强烈依赖于 PEG 水动力半径。这些观察结果与体积排除和吸引力耗竭等空间位阻机制一致。
