Gillery P, Maquart F X, Gattegno L, Randoux A, Cornillot P, Borel J P
Diabetes. 1982 Apr;31(4 Pt 1):371-4. doi: 10.2337/diab.31.4.371.
Red blood cell (RBC) membranes are rich in a glycoconjugate that is extractable in chloroform/methanol solutions (2/1, v/v) and contains several hexoses, such as glucose. Old and young RBC are separated and their respective glycoconjugates are prepared. HbA0 is purified by column chromatography and incubated with solutions of this conjugate. After 24-h incubation, Hb is dialyzed and the amount of glycosylated Hb is measured by a method of column chromatography adapted from Trivelli. A very significant amount of HBAlc is formed when young RBC extracts are incubated: 3.6% of total Hb becomes HBAlc with the extracts, versus 3.2% with free glucose, and only 2.5% for controls. No increase in HbAlc is obtained when extracts of old RBC are incubated. Another difference between the action of the glycoconjugate and free glucose is that the former induces the increase of only the HBAlc fraction, whereas glucose induces the increase of all the minor Hb fractions. The evaluation of glucose contained in the conjugate before and after the glycosylation reaction demonstrates that it is due to an exchange of glucose units from the conjugate to Hb. The reaction is stereospecifically inhibited by p-nitrophenyl-beta-D-glucoside. The nature of the formed HbAlc is demonstrated by isoelectric focusing. A slight increase of HbAlc observed in the incubated controls may be due to an internal migration of some residues of glucose primitively bound to lysyl residues in an unstable form and also to some degree of denaturation during the incubation.
红细胞（RBC）膜富含一种糖缀合物，该糖缀合物可在氯仿/甲醇溶液（2/1，v/v）中提取，且含有多种己糖，如葡萄糖。将老龄和幼龄红细胞分离，并制备它们各自的糖缀合物。通过柱色谱法纯化血红蛋白A0（HbA0），并将其与该糖缀合物的溶液孵育。孵育24小时后，对血红蛋白进行透析，采用改编自特里维利的柱色谱法测定糖化血红蛋白的量。当与幼龄红细胞提取物孵育时，会形成非常显著量的糖化血红蛋白A1c（HBAlc）：提取物使总血红蛋白的3.6%变为HBAlc，而游离葡萄糖使这一比例为3.2%，对照组仅为2.5%。当与老龄红细胞提取物孵育时，HbAlc没有增加。糖缀合物和游离葡萄糖作用的另一个差异在于，前者仅诱导HBAlc组分增加，而葡萄糖诱导所有次要血红蛋白组分增加。对糖基化反应前后缀合物中所含葡萄糖的评估表明，这是由于葡萄糖单元从缀合物交换至血红蛋白所致。该反应受到对硝基苯基-β-D-葡萄糖苷的立体特异性抑制。通过等电聚焦证明了所形成的HbAlc的性质。在孵育的对照组中观察到的HbAlc略有增加，可能是由于一些最初以不稳定形式与赖氨酰残基结合的葡萄糖残基发生内部迁移，以及孵育过程中某种程度的变性所致。
