Structural Insights into the Assembly and Shape of Type III Restriction-Modification (R-M) EcoP15I Complex by Small-Angle X-ray Scattering
Structural Insights into the Assembly and Shape of Type III Restriction-Modification (R-M) EcoP15I Complex by Small-Angle X-ray Scattering
复制标题
DOI：
10.1016/j.jmb.2012.04.026
复制
发表时间：
2012-07-20
影响因子：
5.6
通讯作者：
Aggarwal, Aneel K.
中科院分区：
文献类型：
--
作者：
Gupta, Yogesh K.;Yang, Lin;Aggarwal, Aneel K.
EcoP 15 I是III型限制性内切酶家族的原型，由两个修饰（Mod）亚基组成，然后添加两个（或一个）限制性（Res）亚基。Mod亚基负责DNA识别和甲基化，而Res亚基负责ATP水解和切割。尽管有广泛的生物化学和遗传学研究，仍然没有关于III型限制性内切酶的结构信息。我们在这里提出的EcoP 15 I全酶和Mod（2）亚复合物的小角X射线散射（SAXS）和分析超离心分析。我们发现Mod（2）子复合物具有相对紧凑的形状，其回转半径（R-G）近似为37.4埃，最大尺寸近似为110埃。全酶呈细长的新月形，R-G近似于65.3埃，最大尺寸近似于218埃。从重建的SAXS包膜，我们假设Mod（2）可能停靠在全酶的中间，两端各有一个Res亚基。我们讨论了我们的模型对EcoP 15 I的影响。作用，由此Res亚基可以聚集在一起并在DNA周围形成“滑动夹”。(C)2012爱思唯尔有限公司保留所有权利。
EcoP15I is the prototype of the Type III restriction enzyme family, composed of two modification (Mod) subunits to which two (or one) restriction (Res) subunits are then added. The Mod subunits are responsible for DNA recognition and methylation, while the Res subunits are responsible for ATP hydrolysis and cleavage. Despite extensive biochemical and genetic studies, there is still no structural information on Type III restriction enzymes. We present here small-angle X-ray scattering (SAXS) and analytical ultracentrifugation analysis of the EcoP15I holoenzyme and the Mod(2) subcomplex. We show that the Mod(2) subcomplex has a relatively compact shape with a radius of gyration (R-G) of similar to 37.4 angstrom and a maximal dimension of similar to 110 angstrom. The holoenzyme adopts an elongated crescent shape with an R-G of similar to 65.3 angstrom and a maximal dimension of similar to 218 angstrom. From reconstructed SAXS envelopes, we postulate that Mod(2) is likely docked in the middle of the holoenzyme with a Res subunit at each end. We discuss the implications of our model for EcoP15I. action, whereby the Res subunits may come together and form a "sliding clamp" around the DNA. (C) 2012 Elsevier Ltd. All rights reserved.