Molecular, biochemical and immunological analyses of canine pancreatic DNase I.
Molecular, biochemical and immunological analyses of canine pancreatic DNase I.
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DOI：
10.1093/jb/mvg196
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发表时间：
2003-11
影响因子：
2.7
通讯作者：
Y. Kaneko;H. Takeshita;K. Mogi;T. Nakajima;T. Yasuda;M. Itoi;H. Kuwano;K. Kishi
中科院分区：
文献类型：
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作者：
Y. Kaneko;H. Takeshita;K. Mogi;T. Nakajima;T. Yasuda;M. Itoi;H. Kuwano;K. Kishi
使用三步柱色谱法将来自犬胰腺的 DNase I 纯化 260 倍，达到电泳均质，产率 35%。纯化酶的活性被 20 mM EDTA（一种针对纯化酶和 G-肌动蛋白的特异性抗体）完全抑制。使用 cDNA 末端快速扩增方法，从犬胰腺总 RNA 中构建了编码犬 DNase I 的 1,373 bp cDNA，然后进行测序。成熟的犬 DNase I 蛋白被发现由 262 个氨基酸组成。对 13 种不同犬组织中 DNase I 的调查显示，胰腺中 DNase I 酶活性和基因表达水平最高；因此，犬 DNase I 属于胰腺型。系统发育和序列同一性分析、免疫学特性和 DNase I 组织分布模式的研究表明，犬 DNase I 与人类 DNase I 的关系比与其他哺乳动物 DNase I 的关系更密切。因此，犬 DNase I 被认为是人类 DNase I 研究中最好的替代品之一。
The DNase I from canine pancreas was purified 260-fold to electrophoretic homogeneity with a 35% yield using three-step column chromatography. The activity of the purified enzyme was completely inhibited by 20 mM EDTA, an antibody specific to the purified enzyme and G-actin. A 1,373-bp cDNA encoding canine DNase I was constructed from the total canine pancreatic RNA using a rapid amplification of cDNA ends method, followed by sequencing. The mature canine DNase I protein was found to consist of 262 amino acids. A survey of DNase I in 13 different canine tissues revealed the highest levels of both DNase I enzyme activity and gene expression in the pancreas; therefore, the canine DNase I is of the pancreatic type. Phylogenetic and sequence identity analyses, studies of immunological properties and the tissue-distribution patterns of DNase I indicated that the canine enzyme is more closely related to the human DNase I than to other mammalian DNases I. Therefore, canine DNase I is found to be one of the best substitutes in studies of human DNase I.