HUMAN AND RABBIT PARAOXONASES - PURIFICATION, CLONING, SEQUENCING, MAPPING AND ROLE OF POLYMORPHISM IN ORGANOPHOSPHATE DETOXIFICATION
HUMAN AND RABBIT PARAOXONASES - PURIFICATION, CLONING, SEQUENCING, MAPPING AND ROLE OF POLYMORPHISM IN ORGANOPHOSPHATE DETOXIFICATION
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DOI：
10.1016/0009-2797(93)90023-r
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发表时间：
1993-06-01
影响因子：
5.1
通讯作者：
HUMBERT, R
中科院分区：
文献类型：
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作者：
FURLONG, CE;COSTA, LG;HUMBERT, R
人和家兔对氧磷酶/芳基酯酶。通过色谱和凝胶电泳/等聚焦程序结合活性染色纯化至均匀性。n端和肽序列分析表明保留了分泌信号序列，并允许设计寡核苷酸探针。这些探针被用来分离一个1294 bp的兔对氧磷酶cDNA克隆，该克隆又被用来分离三个人类cDNA克隆。兔和人的蛋白质和cDNA序列比较表明，对氧磷酶具有高度的序列保守性（约85%的同源性），并证实对氧磷酶保留了其信号序列（n端Met除外）。兔的cDNA编码359个氨基酸的蛋白质，人的cDNA编码355个氨基酸的蛋白质。原位杂交证明，正如预期的那样，对氧磷酶基因映射到人类7号染色体的长臂上。192位精氨酸指定高活性对氧磷酶和谷氨酰胺低活性人对氧磷酶。蛋白质水平的变化解释了在一个遗传类中观察到的酶活性的变化。毒性研究表明，通过静脉注射部分纯化的兔对氧磷酶提高大鼠血浆对氧磷酶水平，可以保护动物免受对氧磷和毒死蜱对胆碱酯酶的抑制。保护作用与这两种化合物的相对水解速率有关。
Human and rabbit paraoxonases/arylesterases were.purified to homogeneity by chromatographic and gel electrophoretic/isofocusing procedures coupled with activity stains. N-terminal and peptide sequence analysis suggested retention of the secretion signal sequence and allowed design of oligonucleotide probes. The probes were used to isolate a 1294-bp rabbit paraoxonase cDNA clone, which, in turn, was used to isolate three human cDNA clones. Comparison of rabbit and human protein and cDNA sequences indicated a high degree of sequence conservation (approximately 85% identity) and verified that paraoxonase retains its signal sequence (except for the N-terminal Met). The rabbit cDNA encodes a protein of 359 amino acids and the human a protein of 355 amino acids. In situ hybridization demonstrated, as expected, that the paraoxonase gene maps to the long arm of human chromosome 7. Arginine at position 192 specifies high activity paraoxonase and glutamine low activity human paraoxonase. Variation in protein levels explains the variation of enzyme activity observed within a genetic class. Toxicity studies showed that raising rat plasma paraoxonase levels by i.v. administration of partially purified rabbit paraoxonase protected animals against cholinesterase inhibition by paraoxon and chlorpyrifos oxon. Protection correlated with the relative rates of hydrolysis of these two compounds.