Villanueva Rodrigo A, Guacucano Maritza, Pizarro Jacqueline, Sandino Ana María
Laboratorio de Virología, Departamento de Biología, Facultad de Química y Biología, Universidad de Santiago de Chile, P.O. Box 40, Santiago 33, Chile.
Virus Res. 2005 Sep;112(1-2):132-5. doi: 10.1016/j.virusres.2005.02.011.
Infectious pancreatic necrosis virus (IPNV) is a bi-segmented, dsRNA virus of the Birnaviridae family. The structural protein VP1 has been postulated as the RNA-dependent RNA polymerase (RdRp), but its transcriptional activity has not been unequivocally identified from viral particles. Here, we assayed partially purified IPNV in an in vitro RNA synthesis system. To test the RdRp, dialdehyde-nucleotide analogs were used to covalently inhibit the polymerase-associated activity. Our results showed that dialdehyde-nucleotide analogs completely abrogated IPNV in vitro RNA synthesis. The protein involved in this process was identified as viral VP1, since: (a) after incubation of IPNV with [alpha-(32)P]2',3'-dialdehyde-UTP, labeled VP1 protein was identified and (b) VP1 was unable to bind [alpha-(32)P]GTP when particles were preincubated with 2',3'-dialdehyde-ATP. Thus, within viral particles, inhibition of the transcriptional activity is a result of the binding of 2',3'-dialdehyde-nucleotide analogs to the RdRp, VP1.
传染性胰腺坏死病毒（IPNV）是双RNA病毒科的一种双链RNA病毒。结构蛋白VP1被推测为RNA依赖性RNA聚合酶（RdRp），但其转录活性尚未从病毒颗粒中得到明确鉴定。在此，我们在体外RNA合成系统中检测了部分纯化的IPNV。为了测试RdRp，使用二醛核苷酸类似物共价抑制聚合酶相关活性。我们的结果表明，二醛核苷酸类似物完全消除了IPNV的体外RNA合成。参与此过程的蛋白质被鉴定为病毒VP1，原因如下：（a）用[α-(32)P]2',3'-二醛-UTP孵育IPNV后，鉴定出标记的VP1蛋白；（b）当颗粒与2',3'-二醛-ATP预孵育时，VP1无法结合[α-(32)P]GTP。因此，在病毒颗粒内，转录活性的抑制是2',3'-二醛核苷酸类似物与RdRp（VP1）结合的结果。
