Flexibility of NS5 Methyltransferase-Polymerase Linker Region Is Essential for Dengue Virus Replication

Yongqian Zhao; Tingjin Sherryl Soh; Kitti Wing Ki Chan; Sarah Suet Yin Fung; Kunchithapadam Swaminathan; Siew Pheng Lim; Pei-Yong Shi; Thomas Huber; Julien Lescar; Dahai Luo; Subhash G Vasudevan

doi:10.1128/JVI.01239-15

Flexibility of NS5 Methyltransferase-Polymerase Linker Region Is Essential for Dengue Virus Replication

J Virol. 2015 Oct;89(20):10717-21. doi: 10.1128/JVI.01239-15. Epub 2015 Aug 12.

Authors

Affiliations

¹ Program in Emerging Infectious Diseases, Duke-NUS Graduate Medical School, Singapore NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, Singapore.
² School of Biological Sciences, Nanyang Technological University, Singapore Novartis Institute for Tropical Diseases, Singapore.
³ Program in Emerging Infectious Diseases, Duke-NUS Graduate Medical School, Singapore.
⁴ NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, Singapore Department of Biological Sciences, National University of Singapore, Singapore.
⁵ Novartis Institute for Tropical Diseases, Singapore.
⁶ Research School of Chemistry, Australian National University, Canberra, Australia.
⁷ School of Biological Sciences, Nanyang Technological University, Singapore UPMC UMRS CR7-CNRS ERL 8255-INSERM U1135 Centre d'Immunologie et des Maladies Infectieuses, Centre Hospitalier Universitaire Pitié-Salpêtrière, Faculté de Médecine Pierre et Marie Curie, Paris, France.
⁸ Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore [email protected] [email protected].
⁹ Program in Emerging Infectious Diseases, Duke-NUS Graduate Medical School, Singapore NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, Singapore [email protected] [email protected].

Abstract

We examined the function of the conserved Val/Ile residue within the dengue virus NS5 interdomain linker (residues 263 to 272) by site-directed mutagenesis. Gly substitution or Gly/Pro insertion after the conserved residue increased the linker flexibility and created slightly attenuated viruses. In contrast, Pro substitution abolished virus replication by imposing rigidity in the linker and restricting NS5's conformational plasticity. Our biochemical and reverse genetics experiments demonstrate that NS5 utilizes conformational regulation to achieve optimum viral replication.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Amino Acid Sequence
Animals
Cell Line
Cricetulus
Crystallography, X-Ray
Dengue Virus / chemistry*
Dengue Virus / genetics
Dengue Virus / metabolism
Gene Expression
Humans
Molecular Dynamics Simulation
Molecular Sequence Data
Mutation
Protein Structure, Secondary
Protein Structure, Tertiary
RNA, Viral / chemistry*
RNA, Viral / metabolism
Recombinant Proteins / chemistry
Recombinant Proteins / genetics
Recombinant Proteins / metabolism
Sequence Alignment
Viral Nonstructural Proteins / chemistry*
Viral Nonstructural Proteins / genetics
Viral Nonstructural Proteins / metabolism
Virus Replication / physiology*

Substances

NS5 protein, dengue virus
RNA, Viral
Recombinant Proteins
Viral Nonstructural Proteins