Iourin O, Mattu T S, Mian N, Keir G, Winchester B, Dwek R A, Rudd P M
Department of Biochemistry, University of Oxford, UK.
Glycoconj J. 1996 Dec;13(6):1031-42. doi: 10.1007/BF01053199.
One of the biochemical characteristics of carbohydrate deficient glycoprotein syndromes is the presence of abnormal glycoforms in serum transferrin. Both glycoform heterogeneity and variable site occupancy may, in principle, lead to the generation of a range of glycoforms which contain different numbers of sialic acid residues, and therefore variable amounts of negative charge. Capillary zone electrophoresis was used to resolve the glycoforms of normal human serum transferrin and also of a set of glycoforms which were prepared by digesting the sugars on the intact glycoprotein with sialidase. The sugars on the intact glycoprotein were also modified by a series of exoglycosidase enzymes to produce a series of neutral glycoforms which were-also analysed by capillary zone electrophoresis. The oligosaccharide population of human serum transferrin was analysed by a series of mixed exoglycosidase digests on the released glycan pool and quantified using a novel HPLC strategy. Transferrin was isolated from carbohydrate deficient glycoprotein syndromes type I serum and both the intact glycoforms and released sugars were resolved and quantified. The data presented here confirm the presence of a hexa-, penta- and tetra-sialoforms of human serum transferrin in both normal and carbohydrate deficient glycoprotein syndrome type I serum samples. Consistent with previous reports carbohydrate deficient glycoprotein syndrome type I transferrin also contained a di-sialoform, representing a glycoform in which one of the two N-glycosylation sites is unoccupied, and a non-glycosylated form where both remain unoccupied. This study demonstrates that capillary zone electrophoresis can be used to resolve quantitatively both sialylated and neutral complex type glycoforms, suggesting a rapid diagnostic test for the carbohydrate deficient glycoprotein syndromes group of diseases.
碳水化合物缺乏糖蛋白综合征的生化特征之一是血清转铁蛋白中存在异常糖型。原则上，糖型异质性和可变位点占据都可能导致一系列含有不同数量唾液酸残基的糖型的产生，因此带有可变数量的负电荷。采用毛细管区带电泳来分离正常人血清转铁蛋白的糖型以及一组通过用唾液酸酶消化完整糖蛋白上的糖而制备的糖型。完整糖蛋白上的糖也通过一系列外切糖苷酶进行修饰，以产生一系列中性糖型，这些中性糖型也通过毛细管区带电泳进行分析。通过对释放的聚糖池进行一系列混合外切糖苷酶消化来分析人血清转铁蛋白的寡糖群体，并使用一种新型高效液相色谱策略进行定量。从I型碳水化合物缺乏糖蛋白综合征血清中分离转铁蛋白，并对完整糖型和释放的糖进行分离和定量。此处呈现的数据证实，在正常人和I型碳水化合物缺乏糖蛋白综合征血清样本中均存在人血清转铁蛋白的六唾液酸型、五唾液酸型和四唾液酸型。与先前的报道一致，I型碳水化合物缺乏糖蛋白综合征转铁蛋白还包含一种二唾液酸型，代表一种其中两个N - 糖基化位点之一未被占据的糖型，以及一种两个位点均未被占据的非糖基化形式。本研究表明，毛细管区带电泳可用于定量分离唾液酸化和中性复合型糖型，这表明可对碳水化合物缺乏糖蛋白综合征组疾病进行快速诊断测试。
