Analysis of a Structural Homology Model of the 2'-O-ribose Methyltransferase Domain Within the Vesicular Stomatitis Virus L Protein

Overview

Journal Virology

Specialty Microbiology

Date 2008 Oct 14

PMID 18848710

Citations 15

Authors

Summer E Galloway

Paul E Richardson

Gail W Wertz

Affiliations

Soon will be listed here.

Abstract

The large (L) proteins of non-segmented negative stranded (NNS) RNA viruses contain the core RNA dependent RNA polymerase activity for RNA replication and transcription as well as the activities for polyadenylating and capping the mRNA transcripts and for methylating the cap structures. There is currently no structural information available for these large multi-functional proteins. Phylogenetic analyses have led to the division of the L protein primary structure into six functional domains of high conservation that are linked by variable regions. The studies in this report investigate the role of specific amino acids within domain VI of the VSV L protein, which contains a 2'-O-ribose methyltransferase (MTase) domain. We generated a structural homology model of residues 1644-1842 within domain VI based on the crystal structure determined for the known 2'-O-ribose MTase of E. coli, RrmJ. The information generated by this homology model directed us to residues structurally important for MTase activity and SAM binding. Selected residues were analyzed by site-specific mutagenesis and the mutant L proteins were assayed for their effects on RNA synthesis and cap methylation. The goal of this study was to functionally test the model in order to gain insight into the structural constraints of this region of the L protein. The data presented here revealed specific mutations that affect transcription, replication, and 5' cap methylation, many of which resulted in polymerases temperature sensitive for RNA synthesis.

Citing Articles

Mechanisms of anti-vesicular stomatitis virus activity of deazaneplanocin and its 3-brominated analogs.

Gibbons J, Khadka S, Williams C, Wang L, Schneller S, Liu C Antiviral Res. 2021; 191:105088.

PMID: 34019950 PMC: 8244617. DOI: 10.1016/j.antiviral.2021.105088.

Evidence for N⁷ guanine methyl transferase activity encoded within the modular domain of RNA-dependent RNA polymerase L of a Morbillivirus.

Gopinath M, Shaila M Virus Genes. 2015; 51(3):356-60.

PMID: 26446666 PMC: 7088885. DOI: 10.1007/s11262-015-1252-3.

A role for H/ACA and C/D small nucleolar RNAs in viral replication.

Murray J, Sheng J, Rubin D Mol Biotechnol. 2014; 56(5):429-37.

PMID: 24477674 PMC: 7090452. DOI: 10.1007/s12033-013-9730-0.

Newly identified phosphorylation site in the vesicular stomatitis virus P protein is required for viral RNA synthesis.

Mondal A, Victor K, Pudupakam R, Lyons C, Wertz G J Virol. 2013; 88(3):1461-72.

PMID: 24257610 PMC: 3911598. DOI: 10.1128/JVI.02384-13.

Second-site mutations selected in transcriptional regulatory sequences compensate for engineered mutations in the vesicular stomatitis virus nucleocapsid protein.

Harouaka D, Wertz G J Virol. 2012; 86(20):11266-75.

PMID: 22875970 PMC: 3457138. DOI: 10.1128/JVI.01238-12.

References

Ray D, Shah A, Tilgner M, Guo Y, Zhao Y, Dong H . West Nile virus 5'-cap structure is formed by sequential guanine N-7 and ribose 2'-O methylations by nonstructural protein 5. J Virol. 2006; 80(17):8362-70. PMC: 1563844. DOI: 10.1128/JVI.00814-06. View

Grdzelishvili V, Smallwood S, Tower D, Hall R, Hunt D, Moyer S . Identification of a new region in the vesicular stomatitis virus L polymerase protein which is essential for mRNA cap methylation. Virology. 2006; 350(2):394-405. DOI: 10.1016/j.virol.2006.02.021. View

Zhou Y, Ray D, Zhao Y, Dong H, Ren S, Li Z . Structure and function of flavivirus NS5 methyltransferase. J Virol. 2007; 81(8):3891-903. PMC: 1866096. DOI: 10.1128/JVI.02704-06. View

Malet H, Egloff M, Selisko B, Butcher R, Wright P, Roberts M . Crystal structure of the RNA polymerase domain of the West Nile virus non-structural protein 5. J Biol Chem. 2007; 282(14):10678-89. DOI: 10.1074/jbc.M607273200. View

Li J, Rahmeh A, Morelli M, Whelan S . A conserved motif in region v of the large polymerase proteins of nonsegmented negative-sense RNA viruses that is essential for mRNA capping. J Virol. 2007; 82(2):775-84. PMC: 2224588. DOI: 10.1128/JVI.02107-07. View

Dong H, Ren S, Zhang B, Zhou Y, Puig-Basagoiti F, Li H . West Nile virus methyltransferase catalyzes two methylations of the viral RNA cap through a substrate-repositioning mechanism. J Virol. 2008; 82(9):4295-307. PMC: 2293060. DOI: 10.1128/JVI.02202-07. View

Feller J, Smallwood S, Horikami S, Moyer S . Mutations in conserved domains IV and VI of the large (L) subunit of the sendai virus RNA polymerase give a spectrum of defective RNA synthesis phenotypes. Virology. 2001; 269(2):426-39. DOI: 10.1006/viro.2000.0234. View

Bugl H, Fauman E, Staker B, Zheng F, Kushner S, Saper M . RNA methylation under heat shock control. Mol Cell. 2000; 6(2):349-60. DOI: 10.1016/s1097-2765(00)00035-6. View

Green T, MacPherson S, Qiu S, Lebowitz J, Wertz G, Luo M . Study of the assembly of vesicular stomatitis virus N protein: role of the P protein. J Virol. 2000; 74(20):9515-24. PMC: 112381. DOI: 10.1128/jvi.74.20.9515-9524.2000. View

10.

Furuichi Y, Shatkin A . Viral and cellular mRNA capping: past and prospects. Adv Virus Res. 2000; 55:135-84. PMC: 7131690. DOI: 10.1016/s0065-3527(00)55003-9. View

11.

Cortese C, Feller J, Moyer S . Mutations in domain V of the Sendai virus L polymerase protein uncouple transcription and replication and differentially affect replication in vitro and in vivo. Virology. 2000; 277(2):387-96. DOI: 10.1006/viro.2000.0615. View

12.

Holmes D, Moyer S . The phosphoprotein (P) binding site resides in the N terminus of the L polymerase subunit of sendai virus. J Virol. 2002; 76(6):3078-83. PMC: 135992. DOI: 10.1128/jvi.76.6.3078-3083.2002. View

13.

Bujnicki J, Rychlewski L . In silico identification, structure prediction and phylogenetic analysis of the 2'-O-ribose (cap 1) methyltransferase domain in the large structural protein of ssRNA negative-strand viruses. Protein Eng. 2002; 15(2):101-8. DOI: 10.1093/protein/15.2.101. View

14.

Egloff M, Benarroch D, Selisko B, Romette J, Canard B . An RNA cap (nucleoside-2'-O-)-methyltransferase in the flavivirus RNA polymerase NS5: crystal structure and functional characterization. EMBO J. 2002; 21(11):2757-68. PMC: 125380. DOI: 10.1093/emboj/21.11.2757. View

15.

Wertz G, Moudy R, Ball L . Adding genes to the RNA genome of vesicular stomatitis virus: positional effects on stability of expression. J Virol. 2002; 76(15):7642-50. PMC: 136382. DOI: 10.1128/jvi.76.15.7642-7650.2002. View

16.

Hager J, Staker B, Bugl H, Jakob U . Active site in RrmJ, a heat shock-induced methyltransferase. J Biol Chem. 2002; 277(44):41978-86. DOI: 10.1074/jbc.M205423200. View

17.

Cevik B, Smallwood S, Moyer S . The L-L oligomerization domain resides at the very N-terminus of the sendai virus L RNA polymerase protein. Virology. 2003; 313(2):525-36. DOI: 10.1016/s0042-6822(03)00342-8. View

18.

Fabrega C, Hausmann S, Shen V, Shuman S, Lima C . Structure and mechanism of mRNA cap (guanine-N7) methyltransferase. Mol Cell. 2004; 13(1):77-89. DOI: 10.1016/s1097-2765(03)00522-7. View

19.

Rhodes D, Moyer S, Banerjee A . In vitro synthesis of methylated messenger RNA by the virion-associated RNA polymerase of vesicular stomatitis virus. Cell. 1974; 3(4):327-33. DOI: 10.1016/0092-8674(74)90046-4. View

20.

Abraham G, Rhodes D, Banerjee A . Novel initiation of RNA synthesis in vitro by vesicular stomatitis virus. Nature. 1975; 255(5503):37-40. DOI: 10.1038/255037a0. View