A theoretical estimate for nucleotide sugar demand towards Chinese Hamster Ovary cellular glycosylation.
A theoretical estimate for nucleotide sugar demand towards Chinese Hamster Ovary cellular glycosylation.
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DOI：
10.1038/srep28547
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发表时间：
2016-06-27
影响因子：
4.6
通讯作者：
Kontoravdi C
中科院分区：
文献类型：
--
作者：
Del Val IJ;Polizzi KM;Kontoravdi C
糖基化极大地影响许多最畅销的重组治疗性蛋白（rTP）的安全性和有效性。为了确定控制rTP糖基化的最佳细胞培养物补料策略，有必要了解核苷酸糖（NS）如何被消耗到宿主细胞和rTP糖基化。在这里，我们提出了一个理论框架，整合了CHO细胞的糖蛋白质组，N-连接和O-GalNAc糖基化位点的数量对个别宿主细胞蛋白（HCP），和CHO鞘糖脂的碳水化合物含量，以估计对CHO细胞糖基化的NS的需求。我们已经确定了最丰富的N-连接和O-GalNAc CHO糖蛋白，获得了在CHO细胞蛋白质组中N-连接和O-GalNAc糖位点的加权频率，并推导出了NS消耗对CHO细胞糖基化的化学计量系数。通过将获得的化学计量系数与先前报道的CHO细胞的比生长和生产率的数据相结合，我们观察到NS对糖基化的需求是显著的，因此需要更好地理解糖基化对细胞代谢的负担。NS对CHO细胞糖基化的估计需求可用于合理设计确保最佳和一致的rTP糖基化的补料策略。
Glycosylation greatly influences the safety and efficacy of many of the highest-selling recombinant therapeutic proteins (rTPs). In order to define optimal cell culture feeding strategies that control rTP glycosylation, it is necessary to know how nucleotide sugars (NSs) are consumed towards host cell and rTP glycosylation. Here, we present a theoretical framework that integrates the reported glycoproteome of CHO cells, the number of N-linked and O-GalNAc glycosylation sites on individual host cell proteins (HCPs), and the carbohydrate content of CHO glycosphingolipids to estimate the demand of NSs towards CHO cell glycosylation. We have identified the most abundant N-linked and O-GalNAc CHO glycoproteins, obtained the weighted frequency of N-linked and O-GalNAc glycosites across the CHO cell proteome, and have derived stoichiometric coefficients for NS consumption towards CHO cell glycosylation. By combining the obtained stoichiometric coefficients with previously reported data for specific growth and productivity of CHO cells, we observe that the demand of NSs towards glycosylation is significant and, thus, is required to better understand the burden of glycosylation on cellular metabolism. The estimated demand of NSs towards CHO cell glycosylation can be used to rationally design feeding strategies that ensure optimal and consistent rTP glycosylation.