Dalgaard J Z, Garrett R A, Belfort M
Molecular Genetics Program, Wadsworth Center, New York State Department of Health, Albany 12201-0509.
J Biol Chem. 1994 Nov 18;269(46):28885-92.
The archaeal intron in the 23 S rRNA gene of the hyperthermophile Desulfurococcus mobilis has previously been shown to encode a site-specific DNA endonuclease that contains the LAGLIDADG motif. The enzyme, I-DmoI, has been shown to be active in two forms when expressed in vitro, from RNAs representing either the linear (I-DmoIl) or circular (I-DmoIc) intron. In this study we have overexpressed I-DmoIl and I-DmoIc and purified the enzymes from Escherichia coli. The optimal conditions for the enzymatic activity in vitro were determined, and the enzyme was used to delimit the recognition boundary on its DNA substrate (14-20 nucleotides), an intronless 23 S rRNA gene. Despite belonging to the archaeal kingdom, and being the product of a hyperthermophile, I-DmoI shares many properties with LAGLIDADG intron and intein endonucleases in other kingdoms. These results support the view that these phylogenetically diverse enzymes, which function to mobilize the DNA sequences that encode them, share a common ancestry.
嗜热栖热硫化叶菌（Desulfurococcus mobilis）23S rRNA基因中的古菌内含子先前已被证明可编码一种含有LAGLIDADG基序的位点特异性DNA内切酶。该酶I-DmoI在体外表达时，以代表线性（I-DmoIl）或环状（I-DmoIc）内含子的RNA表达时，已显示出两种活性形式。在本研究中，我们过表达了I-DmoIl和I-DmoIc，并从大肠杆菌中纯化了这些酶。确定了体外酶活性的最佳条件，并使用该酶来界定其DNA底物（14 - 20个核苷酸）（一个无内含子的23S rRNA基因）上的识别边界。尽管I-DmoI属于古菌界，并且是嗜热菌的产物，但它与其他界中的LAGLIDADG内含子和内含肽内切酶具有许多共同特性。这些结果支持了这样一种观点，即这些在系统发育上不同的酶，其功能是移动编码它们的DNA序列，有着共同的祖先。
