Choosing the right sugar: How polymerases select a nucleotide substrate
Choosing the right sugar: How polymerases select a nucleotide substrate
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DOI：
10.1073/pnas.94.5.1619
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发表时间：
1997-03-04
影响因子：
11.1
通讯作者：
Joyce, CM
中科院分区：
文献类型：
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作者：
Joyce, CM
快速浏览一下最近的科学文献可能会给人一种印象，即核酸聚合酶正在遭受身份危机。过去的几年里，我们得到了“E”。大肠杆菌DNA聚合酶I作为逆转录酶“（1），”突变T7 RNA聚合酶作为DNA聚合酶“（2），以及在最近一期的《Proceedings》中，”赋予DNA聚合酶RNA聚合酶活性：逆转录酶中的单个残基控制底物选择“（3）。由于聚合酶传统上根据其底物特异性[使用DNA或RNA模板和脱氧核糖核苷酸（dNTP）或核糖核苷酸（rNTP）]分为四类，因此类别之间的区别模糊可能会给人一种重要的生物屏障已被突破的印象。这不是一个正确的看法，这是由早期报告的情况下，野生型聚合酶不严格区分脱氧核糖和核糖底物。因此，DNA聚合酶的模板特异性显示出一系列严格性（4），从对DNA模板具有相当严格要求的聚合酶到在逆转录病毒复制循环期间需要使用RNA和DNA模板的逆转录酶。大肠杆菌DNA聚合酶I（1）的特异性相当宽松，可以认为是这两个极端之间的中间值。以类似的方式，野生型聚合酶对具有“错误”糖结构的核苷酸底物的区分不是绝对的，并且当Mn 2+取代Mg 2+作为金属辅因子时进一步放松（2，5-7）。结构研究支持这样的观点，即聚合酶之间的相似性超越了基于脱氧核糖或核糖模板和核苷酸的使用的任意分类（最近的综述，参见参考文献。第8至10段）。从六种已发表的聚合酶结构（11-19）以及最近的科学会议上提出的少数其他结构中得出的不可避免的结论是，大多数聚合酶属于聚合酶超家族，并且具有密切相关的活性位点，这些活性位点类似地位于聚合酶裂缝内，其形状与半张开的右手的形状进行了比较（图1）。到目前为止，这种概括的唯一例外是哺乳动物DNA聚合酶β，现在认为它更适合于归属于一个相关但不同的核苷酸转移酶家族（22，23）。因此，似乎存在提供活性位点结构以进行磷酰基转移反应的通用聚合酶模块，并且对该模块的细微修饰实现了每种聚合酶类别的底物特异性特征。
A quick glance at the recent scientific literature might give the impression that nucleic acid polymerases are suffering from an identity crisis. The last few years have brought us ‘‘E. coli DNA polymerase I as a reverse transcriptase’’(1),‘‘A mutant T7 RNA polymerase as a DNA polymerase’’(2), and, in a recent issue of the Proceedings,‘‘Conferring RNA polymerase activity to a DNA polymerase: A single residue in reverse transcriptase controls substrate selection’’(3). Because polymerases have traditionally been divided into four classes based on their substrate specificities [the use of DNA or RNA templates and deoxyribonucleotides (dNTPs) or ribonucleotides (rNTPs)], the blurring of the distinction between classes might give the impression that an important biological barrier has been breached. That this is not a correct perception is suggested by earlier reports of circumstances in which wild-type polymerases do not discriminate strictly between deoxyribo and ribo substrates. Thus the template specificity of DNA polymerases shows a range of stringency (4), from polymerases with a fairly strict requirement for a DNA template, to reverse transcriptases, which need to use both RNA and DNA templates during the retroviral replication cycle. The rather relaxed specificity of Escherichia coli DNA polymerase I (1) can be thought of as intermediate between these two extremes. In an analogous way, the discrimination by wild-type polymerases against nucleotide substrates with the ‘‘wrong’’sugar structure is not absolute, and is further relaxed when Mn2+ replaces Mg2+ as the metal cofactor (2, 5–7). Structural studies support the view that the similarities between polymerases transcend arbitrary divisions into classes on the basis of the use of deoxyribo or ribo templates and nucleotides (for recent reviews, see refs. 8–10). The inescapable conclusion from the six published polymerase structures (11–19), together with a handful of additional structures presented at recent scientific meetings, is that the majority of polymerases belong to a polymerase superfamily and have closely related active sites similarly positioned within a polymerase cleft whose shape has been compared with that of a half open right hand (Fig. 1). Thus far, the only exception to this generalization is mammalian DNA polymerase ß, which is now recognized to be more appropriately assigned to a related but distinct family of nucleotidyl transferases (22, 23). Thus it appears that there is a generic polymerase module that provides the active site architecture to carry out the phosphoryl transfer reaction, and that subtle modifications to this module achieve the substrate specificities characteristic of each polymerase class.