USE OF CULTURED LYMPHOBLASTOID-CELLS FOR THE STUDY OF ABNORMAL ENZYMES - MOLECULAR ABNORMALITY OF A PHOSPHOGLYCERATE KINASE VARIANT ASSOCIATED WITH HEMOLYTIC-ANEMIA
USE OF CULTURED LYMPHOBLASTOID-CELLS FOR THE STUDY OF ABNORMAL ENZYMES - MOLECULAR ABNORMALITY OF A PHOSPHOGLYCERATE KINASE VARIANT ASSOCIATED WITH HEMOLYTIC-ANEMIA
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DOI：
10.1073/pnas.78.4.2587
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发表时间：
1981-01-01
期刊：
影响因子：
--
通讯作者：
YOSHIDA, A
中科院分区：
文献类型：
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作者：
FUJII, H;CHEN, SH;YOSHIDA, A
一种磷酸甘油酸激酶（PGKase; ATP：3-磷酸甘油酸1-磷酸转移酶; X染色体连锁）变体PGKase-Tokyo与酶缺乏、非球形细胞溶血性贫血和神经系统紊乱有关。因为没有足够量的患者红细胞，所以从患者的培养的淋巴母细胞中纯化变体酶至同质。变异淋巴母细胞的酶活性约为正常淋巴母细胞的16%。与正常酶相比，PGKase-Tokyo的比活性较低（反向反应中为正常值的31%，正向反应中为正常值的15%），ATP和3-磷酸甘油酸的Km高于正常值，最佳pH值更酸性，热不稳定性增加。微量肽图谱分析表明，PGKase-Toyko的结构异常是在位置266处从缬氨酸到甲硫氨酸的单个氨基酸取代。使用培养的淋巴母细胞可用于突变酶的结构和功能异常的研究。
A phosphoglycerate kinase (PGKase; ATP:3-phosphoglycerate 1-phosphotransferase; X chromosome-linked) variant, PGKase-Tokyo, is associated with enzyme deficiency, nonspherocytic hemolytic anemia and neurological disturbances. Because a sufficient amount of the patient''s erythrocytes was not available, the variant enzyme was purified to homogeneity from the cultured lymphoblastoid cells of the patient. The enzyme activity of the variant lymphoblastoid cells was about 16% of that of normal lymphoblastoid cells. PGKase-Tokyo, compared to the normal enzyme, had a lower specific activity (31% of normal in the backward reaction and 15% of normal in the forward reaction), higher than normal Km for ATP and 3-phosphoglycerate, a more acidic pH optimum and increased thermal instability. Microscale peptide mapping analysis revealed that the structural abnormality of PGKase-Toyko is a single amino acid substitution from valine to methionine at position 266. The use of the cultured lymphoblastoid cells is useful for the study of structural and functional abnormalities of mutant enzymes.