Plavša Branimir, Rudman Najda, Pociot Flemming, Gornik Olga
Department of Biochemistry and Molecular Biology, University of Zagreb Faculty of Pharmacy and Biochemistry, 10000 Zagreb, Croatia.
Department of Clinical Research, Steno Diabetes Center Copenhagen, 2730 Herlev, Denmark.
Biomedicines. 2025 May 15;13(5):1206. doi: 10.3390/biomedicines13051206.
: -glycosylation is a post-translational modification involving the attachment of oligosaccharides to proteins and is known to influence immunoglobulin G (IgG) effector functions and even antigen binding. IgG contains an evolutionarily conserved -glycosylation site in its fragment crystallizable (Fc) region, while during V-D-J recombination and somatic hypermutation processes it can also obtain -glycosylation sites in its antigen binding fragment (Fab). Our previous study demonstrated altered IgG -glycosylation in children at type 1 diabetes (T1D) onset, with the most prominent changes involving sialylated glycans, hypothesized to mainly come from the Fab region, however, the analytical method used could not distinguish between Fc and Fab. : IgG was isolated from plasma from 118 children with T1D and 98 healthy controls from the Danish Registry of Childhood and Adolescent Diabetes. Isolated IgG was cleaved into Fc and Fab fragments using IdeS enzyme. -glycans were enzymatically released from each fragment, fluorescently labelled with procainamide, and analyzed separately using the UPLC-MS method. Structural annotation of resulting chromatograms was performed using MS/MS. : T1D related -glycosylation changes were more pronounced in the Fab glycans compared to Fc glycans, with five Fab glycans (Man5, Man7, FA2BG1S1, A2G2S2, FA2BG2S1) being significantly altered compared to only one in the Fc region (FA2[3]BG1). Comparing Fc and Fab glycosylation overall reveals stark differences in the types of glycans on each region, with a more diverse and complex repertoire being present in the Fab region. : These findings suggest that -glycosylation changes in early onset T1D predominantly originate from the Fab region, underscoring their potential role in modulating (auto)immunity and highlighting distinct glycosylation patterns between Fc and Fab.
N-糖基化是一种翻译后修饰，涉及寡糖与蛋白质的连接，已知会影响免疫球蛋白G（IgG）的效应功能甚至抗原结合。IgG在其可结晶片段（Fc）区域含有一个进化上保守的N-糖基化位点，而在V-D-J重组和体细胞超突变过程中，它在其抗原结合片段（Fab）中也可获得N-糖基化位点。我们之前的研究表明，1型糖尿病（T1D）发病时儿童的IgG N-糖基化发生改变，最显著的变化涉及唾液酸化聚糖，推测主要来自Fab区域，然而，所使用的分析方法无法区分Fc和Fab。：从丹麦儿童和青少年糖尿病登记处的118名T1D儿童和98名健康对照的血浆中分离IgG。使用IdeS酶将分离的IgG切割成Fc和Fab片段。从每个片段中酶促释放N-聚糖，用普鲁卡因酰胺进行荧光标记，并使用超高效液相色谱-质谱法分别进行分析。使用串联质谱对所得色谱图进行结构注释。：与Fc聚糖相比，T1D相关的N-糖基化变化在Fab聚糖中更为明显，有五种Fab聚糖（Man5、Man7、FA2BG1S1、A2G2S S2、FA2BG2S1）与Fc区域中仅一种（FA2[3]BG1）相比有显著改变。总体比较Fc和Fab糖基化发现每个区域聚糖类型存在明显差异，Fab区域存在更多样化和复杂的聚糖库。：这些发现表明，T1D早期发病时的N-糖基化变化主要源于Fab区域，强调了它们在调节（自身）免疫中的潜在作用，并突出了Fc和Fab之间不同的糖基化模式。
