Different subcellular localization and glycosylation for a functional antibody expressed in Nicotiana tabacum plants and suspension cells
Different subcellular localization and glycosylation for a functional antibody expressed in Nicotiana tabacum plants and suspension cells
复制标题
DOI：
10.1007/s11248-008-9240-1
复制
发表时间：
2009-06-01
影响因子：
3
通讯作者：
Boutry, Marc
中科院分区：
文献类型：
--
作者：
De Muynck, Benoit;Navarre, Catherine;Boutry, Marc
编码治疗性 IgG 抗体 LO-BM2 的重链和轻链的基因以串联或反向会聚方向组装，并在烟草植物和 BY-2 悬浮细胞中表达。串联构建体允许在两种表达系统中更高的表达。从叶细胞间液和 BY-2 培养基中回收的分泌抗体观察到类似的降解模式。降解随着叶龄或培养时间的增加而增加。从叶组织和 BY-2 细胞中纯化的抗体均具有功能。然而，N-糖基化的MS分析显示，从植物中纯化的抗体中，复杂的植物型聚糖是主要类型，而从BY-2细胞中纯化的抗体中，寡甘露糖苷是主要的糖基化类型。 LO-BM2 主要在 BY-2 细胞的内质网中观察到，而在叶细胞中，它主要位于类似于前液泡区室的囊泡中。这些结果以及来自糖苷内切酶 H 研究的结果表明，LO-BM2 从 BY-2 细胞中比从叶细胞中更容易分泌，在叶细胞中，LO-BM2 在高尔基体后区室中积累。
Genes encoding the heavy and light chains of LO-BM2, a therapeutic IgG antibody, were assembled in the tandem or inverted convergent orientation and expressed in Nicotiana tabacum plants and BY-2 suspension cells. The tandem construct allowed higher expression in both expression systems. A similar degradation pattern was observed for the secreted antibody recovered from the leaf intercellular fluid and BY-2 culture medium. Degradation increased with leaf age or culture time. Antibodies purified from leaf tissues and BY-2 cells were both functional. However, MS analysis of the N-glycosylation showed complex plant-type glycans to be the major type in the antibody purified from plants, whereas, oligomannosidic was the major glycosylation type in that purified from BY-2 cells. LO-BM2 was observed mainly in the endoplasmic reticulum of BY-2 cells while, in leaf cells, it was localized mostly to vesicles resembling prevacuolar compartments. These results and those from endoglycosidase H studies suggest that LO-BM2 is secreted from BY-2 cells more readily than from leaf cells where it accumulates in a post-Golgi compartment.