Blast-Induced Neurotrauma Branch, Center for Military Psychiatry and Neuroscience, Walter Reed Army Institute of Research, Silver Spring, MD 20910, USA.
Toxicol In Vitro. 2012 Feb;26(1):182-8. doi: 10.1016/j.tiv.2011.11.006. Epub 2011 Nov 20.
Human prolidase (PROL), which has structural homology to bacterial organophosphate acid anhydrolase that hydrolyze organophosphates and nerve agents has been proposed recently as a potential catalytic bioscavenger. To develop PROL as a catalytic bioscavenger, we evaluated the in vitro hydrolysis efficiency of purified recombinant human PROL against organophosphates and nerve agents. Human liver PROL was purified by chromatographic procedures, whereas recombinant human skin and kidney PROL was expressed in Trichoplusia ni larvae, affinity purified and analyzed by gel electrophoresis. The catalytic efficiency of PROL against diisopropylfluorophosphate (DFP) and nerve agents was evaluated by acetylcholinesterase back-titration assay. Partially purified human liver PROL hydrolyzed DFP and various nerve agents, which was abolished by specific PROL inhibitor showing the specificity of hydrolysis. Both the recombinant human skin and kidney PROL expressed in T. ni larvae showed ∼99% purity and efficiently hydrolyzed DFP and sarin. In contrast to human liver PROL, both skin and kidney PROL showed significantly low hydrolyzing potential against nerve agents soman, tabun and VX. In conclusion, compared to human liver PROL, recombinant human skin and kidney PROL hydrolyze only DFP and sarin showing the substrate specificity of PROL from various tissue sources.
人源脯氨酰寡肽酶（PROL）与细菌有机磷酸酯酶具有结构同源性，后者可水解有机磷酯和神经毒剂，最近被提议作为一种潜在的催化生物清除剂。为了将 PROL 开发为催化生物清除剂，我们评估了纯化的重组人 PROL 对有机磷酯和神经毒剂的体外水解效率。通过色谱程序纯化人肝 PROL，而重组人皮肤和肾 PROL 则在 Trichoplusia ni 幼虫中表达，通过亲和纯化并通过凝胶电泳进行分析。通过乙酰胆碱酯酶反滴定测定法评估 PROL 对二异丙基氟磷酸酯（DFP）和神经毒剂的催化效率。部分纯化的人肝 PROL 水解 DFP 和各种神经毒剂，而特定的 PROL 抑制剂可消除水解，显示出水解的特异性。在 T. ni 幼虫中表达的重组人皮肤和肾 PROL 均显示出约 99%的纯度，并有效地水解 DFP 和沙林。与人肝 PROL 不同，皮肤和肾 PROL 对神经毒剂梭曼、塔崩和 VX 的水解潜力明显较低。总之，与肝 PROL 相比，重组人皮肤和肾 PROL 仅水解 DFP 和沙林，显示出不同组织来源的 PROL 的底物特异性。
