The integrase of the long terminal repeat-retrotransposon Tf1 has a chromodomain that modulates integrase activities
The integrase of the long terminal repeat-retrotransposon Tf1 has a chromodomain that modulates integrase activities
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DOI：
10.1074/jbc.m506363200
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发表时间：
2005-11-25
影响因子：
4.8
通讯作者：
Levin, HL
中科院分区：
文献类型：
--
作者：
Hizi, A;Levin, HL
多种蛋白质中的染色体结构域通过直接与组蛋白H3、DNA或RNA相互作用来介导异染色质的形成。长末端重复序列（LTR）反转录转座子的一个不同的家庭拥有在其整合酶（IN）的染色域，这表明染色域可能控制整合。粟酒裂殖酵母的LTR反转录转座子Tf 1具有很高的活性，在其IN的COOH末端具有一个色结构域。为了测试该色域在整合中的作用，测定了具有和不具有色域的重组IN在体外反应中的活性。全长IN具有与寡核苷酸底物的整合活性，所述寡核苷酸底物模拟插入反应和称为崩解的逆反应。逆转录病毒的IN具有称为3'加工的额外活性，其必须在插入发生之前从病毒cDNA的3'末端去除2-3个核苷酸。由于负链引物位于LTR下游，因此在逆转录过程中添加了这些额外的核苷酸。Tf 1引物的位置表明LTR的3'端没有添加核苷酸。因此，令人惊讶的是，TflIN能够3'切割。这里报道的最出乎意料的结果是，缺乏染色体结构域的IN具有显著更高的活性和显著降低的底物特异性。这些结果表明，酶的活性和特异性都可以通过它们的染色体结构域来调节。
Chromodomains in a variety of proteins mediate the formation of heterochromatin by interacting directly with histone H3, DNA, or RNA. A diverse family of long terminal repeat (LTR)-retrotransposons possesses chromodomains in their integrases (IN), suggesting that the chromodomains may control integration. The LTR-retrotransposon Tf1 of Schizosaccharomyces pombe is highly active and possesses a chromodomain in the COOH terminus of its IN. To test this chromodomain for a role in integration, recombinant INs with and without the chromodomain were assayed for activity in in vitro reactions. The full-length IN had integration activity with oligonucleotide substrates that modeled both the insertion reaction and a reverse reaction known as disintegration. The INs of retroviruses possess an additional activity termed 3' processing that must remove 2-3 nucleotides from the 3' ends of the viral cDNA before insertion can occur. These additional nucleotides are added during reverse transcription because of the position of the minus strand primer downstream of the LTR. The position of the primer for Tf1 suggests no nucleotides are added 3' of the LTR. It was therefore surprising that Tf1 IN was capable of 3' cleavage. The most unexpected result reported here was that the IN lacking the chromodomain had significantly higher activity and substantially reduced substrate specificity. These results reveal that both the activity and specificity of enzymes can be modulated by their chromodomains.