3EVF

Crystal structure of Me7-GpppA complex of yellow fever virus methyltransferase and S-adenosyl-L-homocysteine

Summary for 3EVF

• Entry DOI: 10.2210/pdb3evf/pdb
• Related: 3EVA 3EVB 3EVC 3EVD 3EVE 3EVG
• Descriptor: RNA-directed RNA polymerase NS5, P1-7-METHYLGUANOSINE-P3-ADENOSINE-5',5'-TRIPHOSPHATE, S-ADENOSYL-L-HOMOCYSTEINE, ... (4 entities in total)
• Functional Keywords: yellow fever virus, ns5 methyltransferase, rna cap binding, atp-binding, capsid protein, cleavage on pair of basic residues, endoplasmic reticulum, envelope protein, glycoprotein, helicase, hydrolase, membrane, metal-binding, multifunctional enzyme, nucleotide-binding, nucleotidyltransferase, nucleus, phosphoprotein, protease, ribonucleoprotein, rna replication, rna-binding, rna-directed rna polymerase, secreted, serine protease, transmembrane, viral nucleoprotein, virion, transferase
• Biological source: Yellow fever virus
• Cellular location: Capsid protein C: Virion (Potential). Peptide pr: Secreted. Small envelope protein M: Virion membrane; Multi-pass membrane protein (Potential). Envelope protein E: Virion membrane; Multi- pass membrane protein (Potential). Non-structural protein 1: Secreted. Non-structural protein 2A-alpha: Host endoplasmic reticulum membrane; Multi-pass membrane protein (Potential). Non-structural protein 2A: Host endoplasmic reticulum membrane; Multi-pass membrane protein (Potential). Serine protease subunit NS2B: Host endoplasmic reticulum membrane; Peripheral membrane protein; Cytoplasmic side. Serine protease NS3: Host endoplasmic reticulum membrane; Peripheral membrane protein; Cytoplasmic side (By similarity). Non-structural protein 4A: Host endoplasmic reticulum membrane; Multi-pass membrane protein (By similarity). Non-structural protein 4B: Host endoplasmic reticulum membrane; Multi-pass membrane protein (By similarity). RNA-directed RNA polymerase NS5: Host endoplasmic reticulum membrane; Peripheral membrane protein; Cytoplasmic side (By similarity): P03314
• Total number of polymer chains: 1
• Total formula weight: 33250.10

Authors

Thompson, A.A.,Geiss, B.J.,Peersen, O.B. (deposition date: 2008-10-13, release date: 2009-01-06, Last modification date: 2023-09-06)

Primary citation

Geiss, B.J.,Thompson, A.A.,Andrews, A.J.,Sons, R.L.,Gari, H.H.,Keenan, S.M.,Peersen, O.B.
Analysis of flavivirus NS5 methyltransferase cap binding.
J.Mol.Biol., 385:1643-1654, 2009

PubMed Abstract: The flavivirus 2'-O-nucleoside N-terminal RNA methyltransferase (MTase) enzyme is responsible for methylating the viral RNA cap structure. To increase our understanding of the mechanism of viral RNA cap binding we performed a detailed structural and biochemical characterization of the guanosine cap-binding pocket of the dengue (DEN) and yellow fever (YF) virus MTase enzymes. We solved an improved 2.1 A resolution crystal structure of DEN2 Mtase, new 1.5 A resolution crystal structures of the YF virus MTase domain in apo form, and a new 1.45 A structure in complex with guanosine triphosphate and RNA cap analog. Our structures clarify the previously reported DEN MTase structure, suggest novel protein-cap interactions, and provide a detailed view of guanine specificity. Furthermore, the structures of the DEN and YF proteins are essentially identical, indicating a large degree of structural conservation amongst the flavivirus MTases. Guanosine triphosphate analog competition assays and mutagenesis analysis, performed to analyze the biochemical characteristics of cap binding, determined that the major interaction points are (i) guanine ring via pi-pi stacking with Phe24, N1 hydrogen interaction with the Leu19 backbone carbonyl via a water bridge, and C2 amine interaction with Leu16 and Leu19 backbone carbonyls; (ii) ribose 2' hydroxyl interaction with Lys13 and Asn17; and (iii) alpha-phosphate interactions with Lys28 and Ser215. Based on our mutational and analog studies, the guanine ring and alpha-phosphate interactions provide most of the energy for cap binding, while the combination of the water bridge between the guanine N1 and Leu19 carbonyl and the hydrogen bonds between the C2 amine and Leu16/Leu19 carbonyl groups provide for specific guanine recognition. A detailed model of how the flavivirus MTase protein binds RNA cap structures is presented.

PubMed: 19101564
DOI: 10.1016/j.jmb.2008.11.058

Experimental method

X-RAY DIFFRACTION (1.45 Å)