Abstract：
The presence of α2,6-sialic acids on the Fc N-glycan provides anti-inflammatory properties to the IgGs through a mechanism that remains unclear. Fc-sialylated IgGs are rare in humans as well as in industrial host cell lines such as Chinese hamster ovary (CHO) cells. Facilitated access to well-characterized α2,6-sialylated IgGs would help elucidate the mechanism of this intriguing IgG\'s effector function. This study presents a method for the efficient Fc glycan α2,6-sialylation of a wild-type and a F243A IgG1 mutant by transient co-expression with the human α2,6-sialyltransferase 1 (ST6) and β1,4-galactosyltransferase 1 (GT) in CHO cells. Overexpression of ST6 alone only had a moderate effect on the glycoprofiles, whereas GT alone greatly enhanced Fc-galactosylation, but not sialylation. Overexpression of both GT and ST6 was necessary to obtain a glycoprofile dominated by α2,6-sialylated glycans in both antibodies. The wild-type was composed of the G2FS(6)1 glycan (38%) with remaining unsialylated glycans, while the mutant glycoprofile was essentially composed of G2FS(6)1 (25%), G2FS(3,6)2 (16%) and G2FS(6,6)2 (37%). The α2,6-linked sialic acids represented over 85% of all sialic acids in both antibodies. We discuss how the limited sialylation level in the wild-type IgG1 expressed alone or with GT results from the glycan interaction with Fc\'s amino acid residues or from intrinsic galactosyl- and sialyl-transferases substrate specificities.
摘要：
FcN-聚糖上α2,6-唾液酸的存在通过尚不清楚的机制为IgG提供了抗炎特性。Fc-唾液酸化的IgG在人类以及工业宿主细胞系如中国仓鼠卵巢(CHO)细胞中是罕见的。促进获得充分表征的α2,6-唾液酸化IgG将有助于阐明这种有趣的IgG效应子功能的机制。这项研究提出了一种通过与人α2,6-唾液酸转移酶1（ST6）和β1,4-半乳糖基转移酶1（GT）在CHO细胞中瞬时共表达，对野生型和F243AIgG1突变体进行有效的Fc聚糖α2,6-唾液酸化的方法。单独过表达ST6仅对糖谱有中等影响，而单独的GT大大增强了Fc-半乳糖基化，但不是唾液酸化。GT和ST6的过表达对于在两种抗体中获得由α2,6-唾液酸化聚糖主导的糖谱是必要的。野生型由G2FS（6）1聚糖（38％）和剩余的未唾液酸化聚糖组成，而突变糖谱基本上由G2FS(6)1(25%)组成，G2FS（3,6）2（16%）和G2FS（6,6）2（37%）。α2,6-连接的唾液酸占两种抗体中所有唾液酸的85%以上。我们讨论了单独或与GT一起表达的野生型IgG1中有限的唾液酸化水平是如何由聚糖与Fc的氨基酸残基相互作用或由固有的半乳糖和唾液酸转移酶底物特异性引起的。
