Medvedev Alexei, Viswanathan Prasanth, May Jared, Korba Brent
Department of Microbiology and Immunology, Georgetown University Medical Center, Washington, DC 20057, USA.
Department of Microbiology and Immunology, Georgetown University Medical Center, Washington, DC 20057, USA.
Virology. 2017 Mar;503:37-45. doi: 10.1016/j.virol.2017.01.003. Epub 2017 Jan 20.
The VPg protein of human Norovirus (hNoV) is a multi-functional protein essential for virus replication. The un-cleaved viral precursor protein, ProPol (NS5-6) was 100-fold more efficient in catalyzing VPg nucleotidylylation than the mature polymerase (Pol, NS6), suggesting a specific intracellular role for ProPol. Sequential and single-point alanine substitutions revealed that several positively charged amino acids in the N-terminal region of VPg regulate its nucleotidylylation by ProPol. We provide evidence that VPg directly binds NTPs, inhibition of binding inhibits nucleotidylylation, and NTP binding appears to involve the first 13 amino acids of the protein. Substitution of multiple positively charged amino acids within the first 12 amino acids of the N-terminal region inhibits nucleotidylylation without affecting binding. Substitution of only Lys20 abolishes nucleotidylylation, but not NTP binding. These studies indicate that positively charged amino acids in the first 20 amino acids of hNoV VPg regulate its nucleotidylylation though several potential mechanisms.
人诺如病毒（hNoV）的VPg蛋白是一种对病毒复制至关重要的多功能蛋白。未切割的病毒前体蛋白ProPol（NS5-6）催化VPg核苷酸化的效率比成熟聚合酶（Pol，NS6）高100倍，这表明ProPol在细胞内具有特定作用。序列和单点丙氨酸取代显示，VPg N端区域的几个带正电荷的氨基酸通过ProPol调节其核苷酸化。我们提供的证据表明，VPg直接结合NTPs，结合的抑制会抑制核苷酸化，并且NTP结合似乎涉及该蛋白的前13个氨基酸。N端区域前12个氨基酸内多个带正电荷氨基酸的取代会抑制核苷酸化，但不影响结合。仅赖氨酸20的取代会消除核苷酸化，但不影响NTP结合。这些研究表明，hNoV VPg前20个氨基酸中的带正电荷氨基酸通过几种潜在机制调节其核苷酸化。
