Cottreau D, Boivin P, Kahn A, Milani A, Marie J
Biochimie. 1975;57(3):325-35. doi: 10.1016/s0300-9084(75)80308-7.
Human granulocyte 6 phosphogluconate dehydrogenase has been totally purified from a single patient with chronic granulocytic leukaemia. 48 mg of protein, of specific activity 20 IU per mg of protein, have been obtained in the course of three different steps only. The overall yield was 30 p. cent and the purification was 100 folds. Purified 6 phosphogluconate dehydrogenase was homogeneous when tested in acrylamide and acrylamide SDS gel electrophoresis or in immunodiffusion. The enzyme was immunologically identical in red blood cells, blood platelets and normal leukocytes. The fixation of both substrates, NADP-+ and 6 phosphogluconate, seemed to proceed through a non ordered mechanism. NADPH was an inhibitor strictly competitive with respect to NADP-+ and non competitive with respect to 6 phosphogluconate. 2-3 Diphosphoglycerate seemed to be able to bind on both the fixation sites of NADP-+ and 6 phosphogluconate. The inhibition by ATP was competitive with 6 phosphogluconate and non competitive with NADP-+. 6 phosphogluconate dehydrogenase was inactivated by SH reagents and was partially protected against this inactivation by both substrates. Both substrates protected the enzyme against thermal inactivation. The influence of ionic strength, pH and ions have been studied, and the results have been compared to those reported by other authors for erythrocyte enzyme.
人粒细胞6-磷酸葡萄糖酸脱氢酶已从一名慢性粒细胞白血病患者中完全纯化出来。仅通过三个不同步骤就获得了48毫克蛋白质，其比活性为每毫克蛋白质20国际单位。总产率为30%，纯化倍数为100倍。在丙烯酰胺和丙烯酰胺SDS凝胶电泳或免疫扩散试验中，纯化后的6-磷酸葡萄糖酸脱氢酶是均一的。该酶在红细胞、血小板和正常白细胞中在免疫学上是相同的。两种底物NADP⁺和6-磷酸葡萄糖酸的结合似乎通过无序机制进行。NADPH是对NADP⁺的严格竞争性抑制剂，对6-磷酸葡萄糖酸是非竞争性抑制剂。2,3-二磷酸甘油酸似乎能够结合在NADP⁺和6-磷酸葡萄糖酸的结合位点上。ATP的抑制作用对6-磷酸葡萄糖酸是竞争性的，对NADP⁺是非竞争性的。6-磷酸葡萄糖酸脱氢酶被巯基试剂灭活，两种底物都能部分保护其免受这种灭活。两种底物都能保护该酶免受热灭活。研究了离子强度、pH值和离子的影响，并将结果与其他作者报道的红细胞酶的结果进行了比较。
