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首页> 外文期刊>Nucleic Acids Research >Characterization of the Type III restriction endonuclease PstII from Providencia stuartii.
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Characterization of the Type III restriction endonuclease PstII from Providencia stuartii.

机译：来自斯氏普罗维登斯氏菌的III型限制性核酸内切酶PstII的表征。

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摘要

A new Type III restriction endonuclease designated PstII has been purified from Providencia stuartii. PstII recognizes the hexanucleotide sequence 5'-CTGATG(N)(25-26/27-28)-3'. Endonuclease activity requires a substrate with two copies of the recognition site in head-to-head repeat and is dependent on a low level of ATP hydrolysis ( approximately 40 ATP/site/min). Cleavage occurs at just one of the two sites and results in a staggered cut 25-26 nt downstream of the top strand sequence to generate a two base 5'-protruding end. Methylation of the site occurs on one strand only at the first adenine of 5'-CATCAG-3'. Therefore, PstII has characteristic Type III restriction enzyme activity as exemplified by EcoPI or EcoP15I. Moreover, sequence asymmetry of the PstII recognition site in the T7 genome acts as an historical imprint of Type III restriction activity in vivo. In contrast to other Type I and III enzymes, PstII has a more relaxed nucleotide specificity and can cut DNA with GTP and CTP (but not UTP).We also demonstrate that PstII and EcoP15I cannot interact and cleave a DNA substrate suggesting that Type III enzymes must make specific protein-protein contacts to activate endonuclease activity.
机译：一种新的Ⅲ型限制性核酸内切酶命名为PstII，已从Providencia stuartii纯化。 PstII识别六核苷酸序列5'-CTGATG（N）（25-26 / 27-28）-3'。核酸内切酶活性要求底物在头对头重复中具有两个拷贝的识别位点，并且依赖于低水平的ATP水解（大约40 ATP /位点/分钟）。切割仅发生在两个位点之一，并导致在顶部链序列的下游交错切割25-26nt，从而产生两个碱基的5'突出末端。该位点的甲基化仅在5'-CATCAG-3'的第一个腺嘌呤上发生在一条链上。因此，PstII具有特征性的III型限制性酶活性，如EcoPI或EcoP15I所示。而且，T7基因组中PstII识别位点的序列不对称性是体内III型限制活性的历史印记。与其他I型和III型酶相比，PstII具有更宽松的核苷酸特异性，可以用GTP和CTP切割DNA（但不能用UTP切割）。必须使特定的蛋白质与蛋白质接触以激活核酸内切酶活性。

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