Williams D G
Biochem J. 1983 Mar 1;209(3):817-29. doi: 10.1042/bj2090817.
Neurotoxic esterase activity was measured in homogenates of human placenta and hen brain, spinal cord, liver, kidney and spleen. The activity in liver comprised less than 20% of the Paraoxon-resistant esterases, but in the other tissues neurotoxic esterase accounted for over 50%. The same tissues were labelled with [3H]di-isopropyl phosphorofluoridate, and any isopropyl group transferred on to protein during 'aging' of the labelled enzymes (alkali-volatilizable tritium) was measured. No Paraoxon-sensitive labelled sites were found to age in this way in any tissue. In brain, the Paraoxon-resistant alkali-volatilizable-tritium-labelled sites correlated with the number of neurotoxic esterase labelled sites, indicating that 'aging' and isopropyl group transfer were 100% efficient. The site receiving the transferred isopropyl group was characterized by analysing the distribution of radiolabelled proteins on gel-filtration chromatography in the presence of SDS. In particulate preparations from each tissue, the protein-bound alkali-volatilizable tritium (transferred isopropyl group) was attached to a polypeptide of Mr 178 000. This same polypeptide also bore the isopropyl-phosphoryl group of neurotoxic esterase, indicating that aging of neurotoxic esterase is an intramolecular group transfer. The apparent turnover number for the enzyme (average 1.6 X 10(5) min-1) was approximately the same in each hen tissue, confirming that closely similar enzymes were present in brain, spinal cord, liver and spleen. The apparent turnover for the human enzyme was 1.8-fold higher than that for the hen enzyme. The concentration of the neurotoxic esterase phosphorylated subunit in brain, spinal cord, spleen, placenta and liver was 14.6, 3.8, 7.4, 3.3 and 3.8 pmol/g of tissue. The evidence indicated that neurotoxic esterase is present in each tissue except kidney, and that isopropyl group transfer on 'aging' occurs on this enzyme only. This process is an intramolecular transfer of the group within the same polypeptide.
在人胎盘以及母鸡的脑、脊髓、肝脏、肾脏和脾脏的匀浆中测量了神经毒性酯酶活性。肝脏中的活性占对氧磷抗性酯酶的比例不到20%,但在其他组织中神经毒性酯酶占比超过50%。用[³H]二异丙基氟磷酸酯标记相同的组织,并测量在标记酶“老化”(碱可挥发的氚)过程中转移到蛋白质上的任何异丙基。在任何组织中均未发现以这种方式老化的对氧磷敏感标记位点。在脑中,对氧磷抗性的碱可挥发氚标记位点与神经毒性酯酶标记位点的数量相关,表明“老化”和异丙基转移效率为100%。通过在SDS存在下分析凝胶过滤色谱上放射性标记蛋白质的分布来表征接受转移异丙基的位点。在每个组织的颗粒制剂中,与蛋白质结合的碱可挥发氚(转移的异丙基)附着在分子量为178 000的多肽上。该相同的多肽还带有神经毒性酯酶的异丙基磷酰基,表明神经毒性酯酶的老化是分子内基团转移。该酶的表观转换数(平均1.6×10⁵ min⁻¹)在每个母鸡组织中大致相同,证实脑、脊髓、肝脏和脾脏中存在非常相似的酶。人酶的表观转换数比母鸡酶高1.8倍。脑、脊髓、脾脏、胎盘和肝脏中神经毒性酯酶磷酸化亚基的浓度分别为14.6、3.8、7.4、3.3和3.8 pmol/g组织。证据表明,除肾脏外,每个组织中都存在神经毒性酯酶,并且只有这种酶在“老化”时发生异丙基转移。这个过程是同一多肽内基团的分子内转移。