Molecular basis for specific viral RNA recognition and 2'-O-ribose methylation by the dengue virus nonstructural protein 5 (NS5)

Yongqian Zhao, Tingjin Sherryl Soh, Siew Pheng Lim, Ka Yan Chung, Kunchithapadam Swaminathan, Subhash G. Vasudevan, Pei-Yong Shi, Julien Lescar, Dahai Luo

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Dengue virus (DENV) causes several hundred million human infections and more than 20,000 deaths annually. Neither an efficacious vaccine conferring immunity against all four circulating serotypes nor specific drugs are currently available to treat this emerging global disease. Capping of the DENV RNA genome is an essential structural modification that protects the RNA from degradation by 5' exoribonucleases, ensures efficient expression of viral proteins, and allows escape from the host innate immune response. The large flavivirus nonstructural protein 5 (NS5) (105 kDa) has RNA methyltransferase activities at its N-terminal region, which is responsible for capping the virus RNA genome. The methyl transfer reactions are thought to occur sequentially using the strictly conserved flavivirus 5'RNA sequence as substrate (GpppAG-RNA), leading to the formation of the 5'RNA cap: G0pppAG-RNA→m7G0pppAG-RNA ("cap-0")→m7G0pppAm2'-O-G-RNA ("cap-1"). To elucidate how viral RNA is specifically recognized and methylated, we determined the crystal structure of a ternary complex between the full-length NS5 protein from dengue virus, an octameric cap-0 viral RNA substrate bearing the authentic DENV genomic sequence (5'-m7G0pppA1G2U3U4G5U6U7-3'), and S-adenosyl-L-homocysteine (SAH), the by-product of the methylation reaction. The structure provides for the first time, to our knowledge, a molecular basis for specific adenosine 2'-O-methylation, rationalizes mutagenesis studies targeting the K61-D146-K180-E216 enzymatic tetrad as well as residues lining the RNA binding groove, and offers previously unidentified mechanistic and evolutionary insights into cap-1 formation by NS5, which underlies innate immunity evasion by flaviviruses.

Original languageEnglish (US)
Pages (from-to)14834-14839
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume112
Issue number48
DOIs
StatePublished - Dec 1 2015

Fingerprint

Dengue Virus
Ribose
Viral RNA
Methylation
Flavivirus
RNA
RNA Caps
Proteins
Innate Immunity
Genome
RNA Viruses
RNA Stability
Methyltransferases
Homocysteine
Viral Proteins
Mutagenesis
Adenosine
Immunity
Vaccines
Infection

Keywords

  • 2'-O-ribose methyltransferase
  • Cap-0 RNA
  • Dengue virus
  • Innate immunity evasion
  • Nonstructural protein 5 methyltransferase-polymerase

ASJC Scopus subject areas

  • General

Cite this

Molecular basis for specific viral RNA recognition and 2'-O-ribose methylation by the dengue virus nonstructural protein 5 (NS5). / Zhao, Yongqian; Soh, Tingjin Sherryl; Lim, Siew Pheng; Chung, Ka Yan; Swaminathan, Kunchithapadam; Vasudevan, Subhash G.; Shi, Pei-Yong; Lescar, Julien; Luo, Dahai.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 112, No. 48, 01.12.2015, p. 14834-14839.

Research output: Contribution to journalArticle

Zhao, Yongqian ; Soh, Tingjin Sherryl ; Lim, Siew Pheng ; Chung, Ka Yan ; Swaminathan, Kunchithapadam ; Vasudevan, Subhash G. ; Shi, Pei-Yong ; Lescar, Julien ; Luo, Dahai. / Molecular basis for specific viral RNA recognition and 2'-O-ribose methylation by the dengue virus nonstructural protein 5 (NS5). In: Proceedings of the National Academy of Sciences of the United States of America. 2015 ; Vol. 112, No. 48. pp. 14834-14839.
@article{534f35f0620f4004a4639d357b3bbe33,
title = "Molecular basis for specific viral RNA recognition and 2'-O-ribose methylation by the dengue virus nonstructural protein 5 (NS5)",
abstract = "Dengue virus (DENV) causes several hundred million human infections and more than 20,000 deaths annually. Neither an efficacious vaccine conferring immunity against all four circulating serotypes nor specific drugs are currently available to treat this emerging global disease. Capping of the DENV RNA genome is an essential structural modification that protects the RNA from degradation by 5' exoribonucleases, ensures efficient expression of viral proteins, and allows escape from the host innate immune response. The large flavivirus nonstructural protein 5 (NS5) (105 kDa) has RNA methyltransferase activities at its N-terminal region, which is responsible for capping the virus RNA genome. The methyl transfer reactions are thought to occur sequentially using the strictly conserved flavivirus 5'RNA sequence as substrate (GpppAG-RNA), leading to the formation of the 5'RNA cap: G0pppAG-RNA→m7G0pppAG-RNA ({"}cap-0{"})→m7G0pppAm2'-O-G-RNA ({"}cap-1{"}). To elucidate how viral RNA is specifically recognized and methylated, we determined the crystal structure of a ternary complex between the full-length NS5 protein from dengue virus, an octameric cap-0 viral RNA substrate bearing the authentic DENV genomic sequence (5'-m7G0pppA1G2U3U4G5U6U7-3'), and S-adenosyl-L-homocysteine (SAH), the by-product of the methylation reaction. The structure provides for the first time, to our knowledge, a molecular basis for specific adenosine 2'-O-methylation, rationalizes mutagenesis studies targeting the K61-D146-K180-E216 enzymatic tetrad as well as residues lining the RNA binding groove, and offers previously unidentified mechanistic and evolutionary insights into cap-1 formation by NS5, which underlies innate immunity evasion by flaviviruses.",
keywords = "2'-O-ribose methyltransferase, Cap-0 RNA, Dengue virus, Innate immunity evasion, Nonstructural protein 5 methyltransferase-polymerase",
author = "Yongqian Zhao and Soh, {Tingjin Sherryl} and Lim, {Siew Pheng} and Chung, {Ka Yan} and Kunchithapadam Swaminathan and Vasudevan, {Subhash G.} and Pei-Yong Shi and Julien Lescar and Dahai Luo",
year = "2015",
month = "12",
day = "1",
doi = "10.1073/pnas.1514978112",
language = "English (US)",
volume = "112",
pages = "14834--14839",
journal = "Proceedings of the National Academy of Sciences of the United States of America",
issn = "0027-8424",
number = "48",

}

TY - JOUR

T1 - Molecular basis for specific viral RNA recognition and 2'-O-ribose methylation by the dengue virus nonstructural protein 5 (NS5)

AU - Zhao, Yongqian

AU - Soh, Tingjin Sherryl

AU - Lim, Siew Pheng

AU - Chung, Ka Yan

AU - Swaminathan, Kunchithapadam

AU - Vasudevan, Subhash G.

AU - Shi, Pei-Yong

AU - Lescar, Julien

AU - Luo, Dahai

PY - 2015/12/1

Y1 - 2015/12/1

N2 - Dengue virus (DENV) causes several hundred million human infections and more than 20,000 deaths annually. Neither an efficacious vaccine conferring immunity against all four circulating serotypes nor specific drugs are currently available to treat this emerging global disease. Capping of the DENV RNA genome is an essential structural modification that protects the RNA from degradation by 5' exoribonucleases, ensures efficient expression of viral proteins, and allows escape from the host innate immune response. The large flavivirus nonstructural protein 5 (NS5) (105 kDa) has RNA methyltransferase activities at its N-terminal region, which is responsible for capping the virus RNA genome. The methyl transfer reactions are thought to occur sequentially using the strictly conserved flavivirus 5'RNA sequence as substrate (GpppAG-RNA), leading to the formation of the 5'RNA cap: G0pppAG-RNA→m7G0pppAG-RNA ("cap-0")→m7G0pppAm2'-O-G-RNA ("cap-1"). To elucidate how viral RNA is specifically recognized and methylated, we determined the crystal structure of a ternary complex between the full-length NS5 protein from dengue virus, an octameric cap-0 viral RNA substrate bearing the authentic DENV genomic sequence (5'-m7G0pppA1G2U3U4G5U6U7-3'), and S-adenosyl-L-homocysteine (SAH), the by-product of the methylation reaction. The structure provides for the first time, to our knowledge, a molecular basis for specific adenosine 2'-O-methylation, rationalizes mutagenesis studies targeting the K61-D146-K180-E216 enzymatic tetrad as well as residues lining the RNA binding groove, and offers previously unidentified mechanistic and evolutionary insights into cap-1 formation by NS5, which underlies innate immunity evasion by flaviviruses.

AB - Dengue virus (DENV) causes several hundred million human infections and more than 20,000 deaths annually. Neither an efficacious vaccine conferring immunity against all four circulating serotypes nor specific drugs are currently available to treat this emerging global disease. Capping of the DENV RNA genome is an essential structural modification that protects the RNA from degradation by 5' exoribonucleases, ensures efficient expression of viral proteins, and allows escape from the host innate immune response. The large flavivirus nonstructural protein 5 (NS5) (105 kDa) has RNA methyltransferase activities at its N-terminal region, which is responsible for capping the virus RNA genome. The methyl transfer reactions are thought to occur sequentially using the strictly conserved flavivirus 5'RNA sequence as substrate (GpppAG-RNA), leading to the formation of the 5'RNA cap: G0pppAG-RNA→m7G0pppAG-RNA ("cap-0")→m7G0pppAm2'-O-G-RNA ("cap-1"). To elucidate how viral RNA is specifically recognized and methylated, we determined the crystal structure of a ternary complex between the full-length NS5 protein from dengue virus, an octameric cap-0 viral RNA substrate bearing the authentic DENV genomic sequence (5'-m7G0pppA1G2U3U4G5U6U7-3'), and S-adenosyl-L-homocysteine (SAH), the by-product of the methylation reaction. The structure provides for the first time, to our knowledge, a molecular basis for specific adenosine 2'-O-methylation, rationalizes mutagenesis studies targeting the K61-D146-K180-E216 enzymatic tetrad as well as residues lining the RNA binding groove, and offers previously unidentified mechanistic and evolutionary insights into cap-1 formation by NS5, which underlies innate immunity evasion by flaviviruses.

KW - 2'-O-ribose methyltransferase

KW - Cap-0 RNA

KW - Dengue virus

KW - Innate immunity evasion

KW - Nonstructural protein 5 methyltransferase-polymerase

UR - http://www.scopus.com/inward/record.url?scp=84948670351&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84948670351&partnerID=8YFLogxK

U2 - 10.1073/pnas.1514978112

DO - 10.1073/pnas.1514978112

M3 - Article

VL - 112

SP - 14834

EP - 14839

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

SN - 0027-8424

IS - 48

ER -