Hansen T L, Christensen E, Willems J L, Trijbels J M
Clin Chim Acta. 1983 Jun 30;131(1-2):39-44. doi: 10.1016/0009-8981(83)90350-9.
Pyruvate carboxylase activity was investigated in cultured fibroblasts from a patient shown to have hepatic pyruvate carboxylase deficiency. Under standard conditions, the activity in fibroblasts was 50% of controls (p less than 0.001). Kinetic investigations of the enzyme showed abnormal protein linearity with low activity at low protein concentration. Mixture of homogenates from the patient and a control revealed no endogenous inhibitor. Temperature stability of the mutant enzyme was similar to controls. Apparent kinetic constants for the substrates bicarbonate, ATP and pyruvate were in the patient 2.6 mmol/l, 0.08 mmol/l and 0.10 mmol/l compared to 2.1 mmol/l, 0.13 mmol/l and 0.22 mmol/l in controls, respectively. The 50% inhibitory concentration of oxaloacetate was 0.5 mmol/l in controls. However, no inhibitory effect of oxaloacetate was found for pyruvate carboxylase in fibroblasts from the patient. With acetyl-CoA, the apparent activation constant was 0.21 mmol/l in controls and 0.10 mmol/l in the patient, while the Hill coefficients were similar. These results may be explained by a mutation primarily affecting the transcarboxylation site of pyruvate carboxylase from the patient.
对一名被证明患有肝丙酮酸羧化酶缺乏症患者的培养成纤维细胞中的丙酮酸羧化酶活性进行了研究。在标准条件下，成纤维细胞中的活性为对照的50%（p小于0.001）。对该酶的动力学研究表明，在低蛋白浓度下活性较低时，蛋白质呈异常线性。患者匀浆与对照匀浆的混合物未显示内源性抑制剂。突变酶的温度稳定性与对照相似。患者中底物碳酸氢盐、ATP和丙酮酸的表观动力学常数分别为2.6 mmol/L、0.08 mmol/L和0.10 mmol/L，而对照中分别为2.1 mmol/L、0.13 mmol/L和0.22 mmol/L。草酰乙酸在对照中的50%抑制浓度为0.5 mmol/L。然而，在该患者的成纤维细胞中未发现草酰乙酸对丙酮酸羧化酶有抑制作用。对于乙酰辅酶A，对照中的表观激活常数为0.21 mmol/L，患者中的为0.10 mmol/L，而希尔系数相似。这些结果可能是由主要影响该患者丙酮酸羧化酶转羧基位点的突变所解释的。
