当前位置: X-MOL 学术Biochem. Biophys. Res. Commun. › 论文详情
Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)
Establishment of a novel monoclonal antibody against truncated glycoforms of α-dystroglycan lacking matriglycans
Biochemical and Biophysical Research Communications ( IF 3.1 ) Pub Date : 2021-09-21 , DOI: 10.1016/j.bbrc.2021.09.043
Fumiya Yamasaki , Fumiko Umezawa , Tomohiro Sensui , Masashi Anzo , Hirohito Abo , Chu-Wei Kuo , Kay-Hooi Khoo , Koichi Kato , Hirokazu Yagi , Hiroto Kawashima

α-Dystroglycan (α-DG) is a glycoprotein specifically modified with O-mannosyl glycans bearing long polysaccharides, termed matriglycans, which comprise repeating units of glucuronic acid and xylose. The matriglycan is linked to the O-mannosyl glycan core through two ribitol phosphate units that can be replaced with glycerol phosphate (GroP) units synthesized by fukutin and fukutin-related protein that transfer GroP from CDP-Gro. Here, we found that forced expression of the bacterial CDP-Gro synthase, TagD, from Bacillus subtilis could result in the overproduction of CDP-Gro in human colon carcinoma HCT116 cells. Western blot and liquid chromatography-tandem mass spectrometry analyses indicated that α-DG prepared from the TagD-expressing HCT116 cells contained abundant GroP and lacked matriglycans. Using the GroP-containing recombinant α-DG-Fc, we developed a novel monoclonal antibody, termed DG2, that reacts with several truncated glycoforms of α-DG, including GroP-terminated glycoforms lacking matriglycans; we verified the reactivity of DG2 against various types of knockout cells deficient in the biosynthesis of matriglycans. Accordingly, forced expression of TagD in HCT116 cells resulted in the reduction of matriglycans and an increase in DG2 reactivity. Collectively, our results indicate that DG2 could serve as a useful tool to determine tissue distribution and function of α-DG lacking matriglycans under physiological and pathophysiological conditions.



中文翻译:

抗缺乏基质聚糖的α-dystroglycan截短糖型的新型单克隆抗体的建立

α-Dystroglycan (α-DG) 是一种糖蛋白,经O-甘露糖基聚糖特异性修饰,带有长多糖,称为基质聚糖,包含葡萄糖醛酸和木糖的重复单元。基质聚糖通过两个核糖醇磷酸酯单元连接到O-甘露糖基聚糖核心,该磷酸酯单元可以替换为由 fukutin 和 fukutin 相关蛋白合成的磷酸甘油 (GroP) 单元,从 CDP-Gro 转移 GroP。在这里,我们发现来自枯草芽孢杆菌的细菌 CDP-Gro 合酶 TagD 的强制表达可能导致人结肠癌 HCT116 细胞中 CDP-Gro 的过量产生。蛋白质印迹和液相色谱-串联质谱分析表明,从表达 TagD 的 HCT116 细胞制备的 α-DG 含有丰富的 GroP 并且缺乏基质聚糖。使用含有 GroP 的重组 α-DG-Fc,我们开发了一种新型单克隆抗体,称为 DG2,它与 α-DG 的几种截短糖型反应,包括缺乏基质聚糖的 GroP 末端糖型;我们验证了 DG2 对缺乏基质聚糖生物合成的各种类型敲除细胞的反应性。因此,HCT116 细胞中 TagD 的强制表达导致基质聚糖的减少和 DG2 反应性的增加。总的来说,

更新日期:2021-09-27
down
wechat
bug