人脱氧核糖核酸酶γ的cDNA克隆：其基因的染色体定位及重组蛋白的酶学性质

cDNA cloning of human DNase gamma: chromosomal localization of its gene and enzymatic properties of recombinant protein.

作者信息

Shiokawa D, Hirai M, Tanuma S

机构信息

Department of Biochemistry, Faculty of Pharmaceutical Sciences, Science University of Tokyo, Shinjuku-ku, Tokyo, Japan.

出版信息

Apoptosis. 1998 Mar;3(2):89-95. doi: 10.1023/a:1009692807692.

DOI:10.1023/a:1009692807692

PMID:14646506

Abstract

We report here on the nucleotide sequence of the cDNA encoding human DNase gamma, which is a candidate for an apoptotic endonuclease. The cDNA clone isolated from a human spleen cDNA library is composed of a 918 bp open reading frame encoding a 305 amino acid precursor protein for DNase gamma. Northern blot analysis reveals that the expression of a single transcript of 1.5 kb DNase gamma mRNA is detected in the spleen and liver. The chromosomal localization of DNase gamma gene is mapped to chromosome 3 at region p21.1-p14.2 by fluorescence in situ hybridization (FISH). Characterization of thioredoxin-DNase gamma fusion protein (Trx-hDNase gamma) shows that the recombinant protein has a Ca(2+)/Mg(2+)- or Mn(2+)-dependent endonuclease activity that cleaves chromatin DNA to nucleosomal units. The optimum pH is around 7.2. Zn(2+) and aurintricarboxylic acid (ATA) inhibits the activity in dose-dependent manners. These properties are identical to those of purified DNase gamma.

摘要

我们在此报告编码人脱氧核糖核酸酶γ（DNaseγ）的cDNA的核苷酸序列，它是一种凋亡核酸内切酶的候选物。从人脾cDNA文库中分离出的cDNA克隆由一个918 bp的开放阅读框组成，该开放阅读框编码DNaseγ的305个氨基酸的前体蛋白。Northern印迹分析显示，在脾和肝中检测到1.5 kb DNaseγ mRNA的单一转录本的表达。通过荧光原位杂交（FISH）将DNaseγ基因的染色体定位定位于3号染色体的p21.1 - p14.2区域。硫氧还蛋白-DNaseγ融合蛋白（Trx-hDNaseγ）的特性表明，重组蛋白具有Ca(2+)/Mg(2+)-或Mn(2+)-依赖性核酸内切酶活性，可将染色质DNA切割成核小体单位。最适pH约为7.2。Zn(2+)和金精三羧酸（ATA）以剂量依赖性方式抑制该活性。这些特性与纯化的DNaseγ相同。