Lee C K, Wimmer E
Department of Microbiology, School of Medicine, State University of New York, Stony Brook 11794.
Virology. 1988 Oct;166(2):405-14. doi: 10.1016/0042-6822(88)90511-9.
The polypeptide 3CD of many poliovirus strains can be cleaved at two different amino acid pairs. The viral proteinase 3C and the viral polymerase 3D result from cleavage at a Gln-Gly pair by proteinase 3C, whereas cleavage at a Tyr-Gly pair by proteinase 2A yields the alternative products 3C' and 3D'. Specific mutations were introduced into the 3C'/3D' cleavage site in an infectious cDNA clone of poliovirus type 1 (Mahoney) by oligonucleotide-directed mutagenesis in order to investigate the role of 3C' and 3D' in viral proliferation and to obtain information about the cleavage specificity of 2Apro. Substitution of a threonine residue by an alanine residue at position -2 (P2) of this cleavage site abolished cleavage, whereas substitution of a tyrosine residue by a phenylalanine residue at amino acid position -1 (P1) of the cleavage site did not influence processing. Both mutated cDNA clones produced infectious viruses (T147A and Y148F) on transfection. The phenotypes of the mutant viruses were similar to that of the parental strain. We conclude that (i) 3C' and 3D' are not essential for virus replication, (ii) a Phe-Gly pair at the cleavage site can be cleaved by 2Apro, and (iii) a threonine residue in the P2 position of the cleavage site may be important in substrate recognition by 2Apro.
许多脊髓灰质炎病毒株的多肽3CD可在两个不同的氨基酸对处裂解。病毒蛋白酶3C和病毒聚合酶3D是由蛋白酶3C在谷氨酰胺-甘氨酸对处裂解产生的，而蛋白酶2A在酪氨酸-甘氨酸对处裂解则产生替代产物3C'和3D'。为了研究3C'和3D'在病毒增殖中的作用并获取有关2A蛋白酶裂解特异性的信息，通过寡核苷酸定向诱变将特定突变引入1型脊髓灰质炎病毒（Mahoney）感染性cDNA克隆的3C'/3D'裂解位点。该裂解位点-2（P2）位置的苏氨酸残基被丙氨酸残基取代消除了裂解，而裂解位点氨基酸位置-1（P1）的酪氨酸残基被苯丙氨酸残基取代不影响加工。转染时，两个突变的cDNA克隆都产生了感染性病毒（T147A和Y148F）。突变病毒的表型与亲本毒株相似。我们得出结论：（i）3C'和3D'对病毒复制不是必需的；（ii）裂解位点的苯丙氨酸-甘氨酸对可被2A蛋白酶裂解；（iii）裂解位点P2位置的苏氨酸残基可能在2A蛋白酶识别底物中起重要作用。
