Proteomic Analysis of Chinese Hamster Ovary Cells
Proteomic Analysis of Chinese Hamster Ovary Cells
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DOI：
10.1021/pr300476w
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发表时间：
2012-11-01
影响因子：
4.4
通讯作者：
Betenbaugh, Michael
中科院分区：
文献类型：
--
作者：
Baycin-Hizal, Deniz;Tabb, David L.;Betenbaugh, Michael
为了补充中国仓鼠卵巢（CHO）细胞的最新基因组测序，对CHO细胞进行蛋白质组学分析，包括细胞蛋白质组，分泌组和糖蛋白质组，使用串联质谱法（MS/MS）的多个组分从凝胶电泳，多维液相色谱，和固相提取糖肽（SPEG）。从120个不同的质谱分析产生682 097 MS/MS光谱，93 548独特的肽序列被确定为最多0.02的错误发现率（FDR）。从糖蛋白质组和蛋白质组分析中共鉴定了6164种分组蛋白，代表目前在CHO蛋白质组中鉴定的蛋白质数量增加了8倍。此外，这是第一个专门使用CHO基因组进行的蛋白质组学研究，它提供了更准确的蛋白质鉴定。根据该分析，确定了CHO密码子频率，发现其与人类不同，这将促进人类蛋白质在CHO细胞中的表达。对蛋白质组和mRNA组合数据集的分析表明，多种途径（包括蛋白质加工和细胞凋亡）的富集，但参与类固醇激素和鞘糖脂代谢的蛋白质的消耗。504个检测到的蛋白质包括N-乙酰化修饰，1292个不同的蛋白质被观察到N-糖基化。这第一次大规模的蛋白质组学分析将增强有关CHO重组表达能力的知识基础，并为旨在修饰CHO细胞功能的细胞工程工作提供有用的信息。
To complement the recent genomic sequencing of Chinese hamster ovary (CHO) cells, proteomic analysis was performed on CHO cells including the cellular proteome, secretome, and glycoproteome using tandem mass spectrometry (MS/MS) of multiple fractions obtained from gel electrophoresis, multidimensional liquid chromatography, and solid phase extraction of glycopeptides (SPEG). From the 120 different mass spectrometry analyses generating 682 097 MS/MS spectra, 93 548 unique peptide sequences were identified with at most 0.02 false discovery rate (FDR). A total of 6164 grouped proteins were identified from both glycoproteome and proteome analysis, representing an 8-fold increase in the number of proteins currently identified in the CHO proteome. Furthermore, this is the first proteomic study done using the CHO genome exclusively, which provides for more accurate identification of proteins. From this analysis, the CHO codon frequency was determined and found to be distinct from humans, which will facilitate expression of human proteins in CHO cells. Analysis of the combined proteomic and mRNA data sets indicated the enrichment of a number of pathways including protein processing and apoptosis but depletion of proteins involved in steroid hormone and glycosphingolipid metabolism. Five hundred four of the detected proteins included N-acetylation modifications, and 1292 different proteins were observed to be N-glycosylated. This first large-scale proteomic analysis will enhance the knowledge base about CHO capabilities for recombinant expression and provide information useful in cell engineering efforts aimed at modifying CHO cellular functions.