dUTPase and nucleocapsid polypeptides of the Mason-Pfizer monkey virus form a fusion protein in the virion with homotrimeric organization and low catalytic efficiency
dUTPase and nucleocapsid polypeptides of the Mason-Pfizer monkey virus form a fusion protein in the virion with homotrimeric organization and low catalytic efficiency
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DOI：
10.1074/jbc.m306967200
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发表时间：
2003-10-03
影响因子：
4.8
通讯作者：
Vértessy, BG
中科院分区：
文献类型：
--
作者：
Barabás, O;Rumlová, M;Vértessy, BG
β逆转录病毒在gag和pol之间的pro开放阅读框中编码dUTR，dUTR是DNA代谢和修复的重要因子。逆转录病毒蛋白表达过程中的核糖体移码为核衣壳（NC）和dUTR在Gag-Pro多聚蛋白中的共价连接提供了独特的可能性。通过开发针对原型β逆转录病毒Mason-Pfizer猴病毒dUTR的抗体，我们证明了i）NC-dUTR融合蛋白存在于病毒体和感染细胞内，提供唯一形式的dUTR，和ii）逆转录病毒蛋白酶不在病毒体或体外切割NC-dUTR。我们表明，重组β逆转录病毒NC-dUTPases和dUTPases都是低效的催化剂相比，所有其他dUTPases。动态光散射和凝胶过滤证实，同源三聚体的组织，常见的dUTPases，保留在NC-dUTPases融合蛋白。β逆转录病毒dUTR已经结晶，并且单晶含有同源三聚体。寡核苷酸和Zn 2+结合很好地保留在融合蛋白中，这是通过DNA修复因子dUTR获得功能性核酸结合模块的第一个例子。六核苷酸ACTGCC或八核苷酸（TG）（4）与NC-dUTR的结合可调节酶功能，表明低催化活性可通过适当的定位得到补偿。
Betaretroviruses encode dUTPase, an essential factor in DNA metabolism and repair, in the pro open reading frame located between gag and pol. Ribosomal frame-shifts during expression of retroviral proteins provide a unique possibility for covalent joining of nucleocapsid (NC) and dUTPase within Gag-Pro polyproteins. By developing an antibody against the prototype betaretrovirus Mason-Pfizer monkey virus dUTPase, we demonstrate that i) the NC-dUTPase fusion protein exists both within the virions and infected cells providing the only form of dUTPase, and ii) the retroviral protease does not cleave NC-dUTPase either in the virion or in vitro. We show that recombinant betaretroviral NC-dUTPase and dUTPase are both inefficient catalysts compared with all other dUTPases. Dynamic light scattering and gel filtration confirm that the homotrimeric organization, common among dUTPases, is retained in the NC-dUTPase fusion protein. The betaretroviral dUTPase has been crystallized and single crystals contain homotrimers. Oligonucleotide and Zn2+ binding is well retained in the fusion protein, which is the first example of acquisition of a functional nucleic acid binding module by the DNA repair factor dUTPase. Binding of the hexanucleotide ACTGCC or the octanucleotide (TG)(4) to NC-dUTPase modulates enzymatic function, indicating that the low catalytic activity may be compensated by adequate localization.