Bakker Hans, Rouwendal Gerard J A, Karnoup Anton S, Florack Dion E A, Stoopen Geert M, Helsper Johannes P F G, van Ree Ronald, van Die Irma, Bosch Dirk
Business Unit Bioscience, Plant Research International, Wageningen University and Research Center, Droevendaalsesteeg 1, 6708 PB, Wageningen, The Netherlands.
Proc Natl Acad Sci U S A. 2006 May 16;103(20):7577-82. doi: 10.1073/pnas.0600879103. Epub 2006 May 4.
N-glycosylation of a mAb may have a major impact on its therapeutic merits. Here, we demonstrate that expression of a hybrid enzyme (called xylGalT), consisting of the N-terminal domain of Arabidopsis thaliana xylosyltransferase and the catalytic domain of human beta-1,4-galactosyltransferase I (GalT), in tobacco causes a sharp reduction of N-glycans with potentially immunogenic core-bound xylose (Xyl) and fucose (Fuc) residues as shown by Western blot and MALDI-TOF MS analysis. A radioallergosorbent test inhibition assay with proteins purified from leaves of WT and these transgenic tobacco plants using sera from allergic patients suggests a significant reduction of potential immunogenicity of xylGalT proteins. A mAb purified from leaves of plants expressing xylGalT displayed an N-glycan profile that featured high levels of galactose, undetectable xylose, and a trace of fucose. Hence, a transgenic plant expressing the hybrid GalT might yield more effective and safer monoclonals for therapeutic purposes than WT plants and even transgenic plants expressing the unchanged GalT.
单克隆抗体（mAb）的N-糖基化可能对其治疗效果产生重大影响。在此，我们证明，在烟草中表达一种由拟南芥木糖基转移酶的N端结构域和人β-1,4-半乳糖基转移酶I（GalT）的催化结构域组成的杂合酶（称为xylGalT），会导致N-聚糖显著减少，如蛋白质印迹和基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）分析所示，这些N-聚糖带有潜在免疫原性的核心结合木糖（Xyl）和岩藻糖（Fuc）残基。使用过敏患者血清对从野生型（WT）和这些转基因烟草植物叶片中纯化的蛋白质进行的放射变应原吸附试验抑制分析表明，xylGalT蛋白的潜在免疫原性显著降低。从表达xylGalT的植物叶片中纯化的单克隆抗体显示出一种N-聚糖谱，其特征是半乳糖水平高、木糖不可检测且岩藻糖含量微量。因此，与野生型植物甚至表达未改变的GalT的转基因植物相比，表达杂合GalT的转基因植物可能产生更有效、更安全的用于治疗目的的单克隆抗体。
