Complex recognition site for the group I intron-encoded endonuclease I-SceII.
Complex recognition site for the group I intron-encoded endonuclease I-SceII.
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I 组内含子编码的核酸内切酶 I-SceII 的复杂识别位点。
DOI：
10.1128/mcb.12.2.716-723.1992
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发表时间：
1992
影响因子：
5.3
通讯作者：
Butow,RA
中科院分区：
文献类型：
--
作者：
Wernette,C;Saldanha,R;Smith,D;Ming,D;Perlman,PS;Butow,RA
我们的特点是由双链DNA核酸内切酶，I-SceII，由内含子4a的酵母casealCOX 1基因编码的网站识别的功能。我们确定了36个点突变对含有theSaccharomycescapensisI-SceII位点的天然和合成底物的切割效率的影响。大多数18 bp的I-SceII识别位点的突变是耐受的酶，这些突变位点被切割42和100%之间，以及野生型底物。9个突变体阻断切割至野生型的≤33%，而只有3个点突变G-4→C、G-12→T和G-15→C完全阻断切割。竞争实验表明，这三种底物不被切割，至少部分是因为酶对这些突变DNA的亲和力显著降低。约90%的源自4-bp交错I-SceII切割位点的核苷酸序列的随机化的DNA不被酶切割。I-SceII大约每110 kbp切割一次来自人3号染色体的克隆DNA。在四个随机选择的人类DNA克隆的I-SceII识别位点有56至78%的同一性与酵母线粒体DNA中的18-bp位点，它们被切割至少50%，以及野生型线粒体底物，尽管存在一些取代，个别损害线粒体底物的切割。这些数据的分析表明，一个给定的碱基取代对I-SceII切割的影响可能取决于在其他位置的序列。
We have characterized features of the site recognized by a double-stranded DNA endonuclease, I-SceII, encoded by intron 4a of the yeast mitochondrialCOX1gene. We determined the effects of 36 point mutations on the cleavage efficiency of natural and synthetic substrates containing theSaccharomyces capensisI-SceII site. Most mutations of the 18-bp I-SceII recognition site are tolerated by the enzyme, and those mutant sites are cleaved between 42 and 100% as well as the wild-type substrate is. Nine mutants blocked cleavage to ≤33% of the wild-type, whereas only three point mutations, G-4→C, G-12→T, and G-15→C, block cleavage completely. Competition experiments indicate that these three substrates are not cleaved, at least in part because of a marked reduction in the affinity of the enzyme for those mutant DNAs. About 90% of the DNAs derived from randomization of the nucleotide sequence of the 4-bp staggered I-SceII cleavage site are not cleaved by the enzyme. I-SceII cleaves cloned DNA derived from human chromosome 3 about once every 110 kbp. The I-SceII recognition sites in four randomly chosen human DNA clones have 56 to 78% identity with the 18-bp site in yeast mitochondrial DNA; they are cleaved at least 50% as well as the wild-type mitochondrial substrate despite the presence of some substitutions that individually compromise cleavage of the mitochondrial substrate. Analysis of these data suggests that the effect of a given base substitution on I-SceII cleavage may depend on the sequence at other positions.