2P3Q

Crystal Structure of Dengue Methyltransferase in Complex with GpppG and S-Adenosyl-L-homocysteine

Summary for 2P3Q

• Entry DOI: 10.2210/pdb2p3q/pdb
• Related: 1L9K 2P1D 2P3L 2P3O 2P40 2P41
• Descriptor: type II methyltransferase, SULFATE ION, DIGUANOSINE-5'-TRIPHOSPHATE, ... (5 entities in total)
• Functional Keywords: vizier; viral enzymes involved in replication; dengue virus methyltransferase; structural genomics; marseilles structural genomics program @ afmb; msgp, viral protein, transferase
• Biological source: Dengue virus 2
• Cellular location: Envelope protein E: Virion membrane; Multi- pass membrane protein: Q9WLZ8
• Total number of polymer chains: 1
• Total formula weight: 36146.22

Authors

Egloff, M.P.,Marseilles Structural Genomics Program @ AFMB (MSGP) (deposition date: 2007-03-09, release date: 2007-08-28, Last modification date: 2023-08-30)

Primary citation

Egloff, M.P.,Decroly, E.,Malet, H.,Selisko, B.,Benarroch, D.,Ferron, F.,Canard, B.
Structural and functional analysis of methylation and 5'-RNA sequence requirements of short capped RNAs by the methyltransferase domain of dengue virus NS5
J.Mol.Biol., 372:723-736, 2007

PubMed Abstract: The N-terminal 33 kDa domain of non-structural protein 5 (NS5) of dengue virus (DV), named NS5MTase(DV), is involved in two of four steps required for the formation of the viral mRNA cap (7Me)GpppA(2'OMe), the guanine-N7 and the adenosine-2'O methylation. Its S-adenosyl-l-methionine (AdoMet) dependent 2'O-methyltransferase (MTase) activity has been shown on capped (7Me+/-)GpppAC(n) RNAs. Here we report structural and binding studies using cap analogues and capped RNAs. We have solved five crystal structures at 1.8 A to 2.8 A resolution of NS5MTase(DV) in complex with cap analogues and the co-product of methylation S-adenosyl-l-homocysteine (AdoHcy). The cap analogues can adopt several conformations. The guanosine moiety of all cap analogues occupies a GTP-binding site identified earlier, indicating that GTP and cap share the same binding site. Accordingly, we show that binding of (7Me)GpppAC(4) and (7Me)GpppAC(5) RNAs is inhibited in the presence of GTP, (7Me)GTP and (7Me)GpppA but not by ATP. This particular position of the cap is in accordance with the 2'O-methylation step. A model was generated of a ternary 2'O-methylation complex of NS5MTase(DV), (7Me)GpppA and AdoMet. RNA-binding increased when (7Me+/-)GpppAGC(n-1) starting with the consensus sequence GpppAG, was used instead of (7Me+/-)GpppAC(n). In the NS5MTase(DV)-GpppA complex the cap analogue adopts a folded, stacked conformation uniquely possible when adenine is the first transcribed nucleotide at the 5' end of nascent RNA, as it is the case in all flaviviruses. This conformation cannot be a functional intermediate of methylation, since both the guanine-N7 and adenosine-2'O positions are too far away from AdoMet. We hypothesize that this conformation mimics the reaction product of a yet-to-be-demonstrated guanylyltransferase activity. A putative Flavivirus RNA capping pathway is proposed combining the different steps where the NS5MTase domain is involved.

PubMed: 17686489
DOI: 10.1016/j.jmb.2007.07.005

Experimental method

X-RAY DIFFRACTION (2.75 Å)