Tangential flow filtration for continuous processing of crystallized proteins
Tangential flow filtration for continuous processing of crystallized proteins
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DOI：
10.1016/j.seppur.2024.126311
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发表时间：
2024-01-12
影响因子：
8.6
通讯作者：
Zydney，Andrew L.
中科院分区：
文献类型：
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作者：
Behboudi，Ali;Minervini，Mirko;Zydney，Andrew L.
蛋白质结晶具有在单个加工步骤中提供几乎纯的蛋白质产品的潜力。大多数以前的蛋白质结晶研究都使用离心脱水和洗涤结晶固体，但这种方法很难在连续的生物加工操作中采用。这项研究首次证明了使用中空纤维切向流过滤（TFF）模块可以成功过滤结晶蛋白质。实验进行了与人血清白蛋白（HSA）结晶通过添加三价氯化铈，有和没有聚乙二醇（PEG）。在3wt%PEG的存在下使用4.5mM CeCl 3实现了大于85%的蛋白质产率，其中结晶发生在小于20秒内，这对于连续加工是理想的。PEG的添加对结晶产率的影响最小，但它确实导致形成更大和更致密的晶体以及临界通量的相应增加，临界通量是导致跨膜压力增加最小的通量的最大值（即，最小的结垢）。在最佳条件下进行的过滤实验成功地运行了24小时，跨膜压力保持低于5 kPa，证明了使用连续结晶与切向流过滤纯化高价值蛋白质产品的可行性。
Protein crystallization has the potential to provide nearly pure protein products in a single processing step. Most previous studies of protein crystallization have used centrifugation for dewatering and washing the crystallized solids, but this approach would be difficult to adopt in continuous bioprocessing operations. This study provides the first demonstration that crystallized proteins can be successfully filtered using hollow fiber tangential flow filtration (TFF) modules. Experiments were performed with human serum albumin (HSA) crystallized by the addition of trivalent CeCl3, both with and without polyethylene glycol (PEG). Greater than 85 % protein yield was achieved using 4.5 mM CeCl3in the presence of 3 wt% PEG with the crystallization occurring in less than 20 s, which is ideal for continuous processing. The addition of PEG had minimal effect on the crystallization yield, but it did lead to the formation of larger and denser crystals and a corresponding increase in the critical flux, which is the maximum value of the flux that causes minimal increase in the transmembrane pressure (i.e., minimal fouling). Filtration experiments performed at optimal conditions were successfully run for 24 h with the transmembrane pressure remaining below 5 kPa, demonstrating the feasibility of using continuous crystallization with tangential flow filtration for the purification of high value protein products.