Human butyrylcholinesterase (HuBChE) is a stoichiometric bioscavenger that protects from the toxicity of nerve agents. Non-human primates are suitable models for toxicity studies that cannot be performed in humans. We evaluated the biochemical properties of native macaque (MaBChE) tetramers, compared to recombinant MaBChE monomers, PEGylated recombinant MaBChE tetramers and monomers, and native HuBChE tetramers. Km and kcat values for butyrylthiocholine were independent of subunit assembly status. The Km for all forms of MaBChE was about 70 μM, compared to 13 μM for HuBChE. The kcat was about 100,000 min-1 for MaBChE and 30,000 min-1 for HuBChE. The reversible inhibitor ethopropazine had similar Ki values of 0.05 μM for all MaBChE forms and HuBChE. The bimolecular rate constant, ki, for inhibition by diisopropylfluorophosphate (DFP), an analog of sarin, was 2.2 to 2.5 × 107 M-1 min-1 for all MaBChE forms and for HuBChE. A major difference between MaBChE and HuBChE was the rate of reactivation by 2-PAM. The second order rate constant for reactivation of DFP-inhibited MaBChE by 2-PAM was 1.4 M-1 min-1, but was 380 fold faster for DFP-inhibited HuBChE (kr 531 M-1 min-1). The acyl pocket of MaBChE has Leu285 in place of Pro285 in HuBChE. The reactivation rate of DFP-inhibited HuBChE mutant P285L by 2-PAM was reduced 5.8-fold (kr 92 M-1 min-1) indicating that P285 determines whether 2-PAM binds in an orientation that favors release of diisopropylphosphate. DFP-inhibited MaBChE treated with 0.2 M 2-PAM recovered 10% of its original activity, whereas DFP-inhibited HuBChE recovered 80% activity. It was concluded that the biochemical properties of MaBChE are similar to those of HuBChE except for the reactivation of DFP-inhibited BChE.

中文翻译：

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脯氨酸285是通过2-PAM重新激活有机磷酸酯抑制的人丁酰胆碱酯酶所不可或缺的。

人丁酰胆碱酯酶（HuBChE）是一种化学计量的生物清除剂，可防止神经毒剂的毒性。非人类灵长类动物是无法在人类中进行毒性研究的合适模型。我们评估了天然猕猴（MaBChE）四聚体与重组MaBChE单体，聚乙二醇化的重组MaBChE四聚体和单体以及天然HuBChE四聚体的生化特性。丁酰硫胆碱的Km和kcat值与亚基组装状态无关。所有形式的MaBChE的Km约为70μM，而HuBChE的Km为13μM。对于MaBChE，kcat约为100,000 min-1，对于HuBChE，kcat约为30,000 min-1。对于所有MaBChE形式和HuBChE，可逆抑制剂乙丙嗪的Ki值相似，均为0.05μM。用于抑制二异丙基氟磷酸酯（DFP）的双分子速率常数ki 沙林的一个类似物，对于所有MaBChE形式和HuBChE，为2.2至2.5×107 M-1 min-1。MaBChE和HuBChE之间的主要区别是2-PAM的重新激活速率。2-PAM重新激活DFP抑制的MaBChE的二级速率常数为1.4 M-1 min-1，但DFP抑制的HuBChE的二级速率常数为380倍（kr 531 M-1 min-1）。MaBChE的酰基口袋中的Leu285代替了HuBChE中的Pro285。2-PAM对DFP抑制的HuBChE突变体P285L的再激活率降低了5.8倍（kr 92 M-1 min-1），表明P285确定2-PAM是否以有利于磷酸二异丙酯释放的方向结合。用0.2 M 2-PAM处理的DFP抑制的MaBChE恢复了其原始活性的10％，而DFP抑制的HuBChE恢复了80％的活性。