Holland M J, DiLorenzo A M, Dancis J, Balis M E, Yü T F, Cox R P
J Clin Invest. 1976 Jun;57(6):1600-5. doi: 10.1172/JCI108430.
Discordance between clinical phenotype and the level of a mutant enzyme activity may reflect differences between enzyme function in vivo and that measured by the customary enzyme assays on cell extracts. In the present study, the conversion of hypoxanthine to phosphorylated products was measured in intact skin fibroblasts and in cell extracts from seven patients with mutant hypoxanthine-guanine phosphoribosyltransferase (HPRT) and six control subjects. The patient's phenotypes ranged from asymptomatic hyperuricemia to the Lesch-Nyhan syndrome. Although there was a general correlation between the HPRT activity in cell extracts assayed by the usual methods and the function of the purine salvage pathway in patients, as reflected by urinary oxypurine excretion, there were notable exceptions. A more accurate appraisal of the functioning of the pathway at the cellular level is achieved by measuring the conversion of substrate to product in the intact cell at physiological concentrations of substrates, activators, and product and metabolite inhibitors, and in a physiological ionic environment. In one of the seven patients, the standard enzyme assay indicated normal function, whereas measurements in the intact cell exposed severe dysfunction of the salvage system. In another, the standard assay suggested a severe deficiency not evident in the intact cell or in the patient.
临床表型与突变酶活性水平之间的不一致可能反映了体内酶功能与通过对细胞提取物进行常规酶测定所测得的酶功能之间的差异。在本研究中，对7名患有突变型次黄嘌呤 - 鸟嘌呤磷酸核糖转移酶（HPRT）的患者和6名对照受试者的完整皮肤成纤维细胞及细胞提取物中次黄嘌呤向磷酸化产物的转化进行了测定。患者的表型从无症状高尿酸血症到莱施 - 奈恩综合征不等。尽管通过常规方法测定的细胞提取物中的HPRT活性与患者嘌呤补救途径的功能之间存在总体相关性，这可通过尿中氧嘌呤排泄反映出来，但也有显著例外。通过在底物、激活剂、产物和代谢物抑制剂的生理浓度下以及在生理离子环境中测量完整细胞中底物向产物的转化，可以更准确地评估该途径在细胞水平的功能。在7名患者中的1名患者中，标准酶测定表明功能正常，而在完整细胞中的测量显示补救系统存在严重功能障碍。在另一名患者中，标准测定表明存在严重缺陷，但在完整细胞或患者中并不明显。
