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A Structural Overview of RNA-Dependent RNA Polymerases from the Flaviviridae Family.

Wu J, Liu W, Gong P - Int J Mol Sci (2015)

Bottom Line: Upon the transition to the elongation phase, this priming element needs to undergo currently unresolved conformational rearrangements to accommodate the growth of the template-product RNA duplex.In the genera of Flavivirus and Pestivirus, the polymerase module in the C-terminal part of the RdRP protein may be regulated in cis by the N-terminal region of the same polypeptide.Either being a methyltransferase in Flavivirus or a functionally unclarified module in Pestivirus, this region could play auxiliary roles for the canonical folding and/or the catalysis of the polymerase, through defined intra-molecular interactions.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Chinese Academy of Sciences, No. 44 Xiao Hong Shan, Wuchang District, Wuhan 430071, China. wujiqin2010@163.com.

ABSTRACT
RNA-dependent RNA polymerases (RdRPs) from the Flaviviridae family are representatives of viral polymerases that carry out RNA synthesis through a de novo initiation mechanism. They share a ≈ 600-residue polymerase core that displays a canonical viral RdRP architecture resembling an encircled right hand with palm, fingers, and thumb domains surrounding the active site. Polymerase catalytic motifs A-E in the palm and motifs F/G in the fingers are shared by all viral RdRPs with sequence and/or structural conservations regardless of the mechanism of initiation. Different from RdRPs carrying out primer-dependent initiation, Flaviviridae and other de novo RdRPs utilize a priming element often integrated in the thumb domain to facilitate primer-independent initiation. Upon the transition to the elongation phase, this priming element needs to undergo currently unresolved conformational rearrangements to accommodate the growth of the template-product RNA duplex. In the genera of Flavivirus and Pestivirus, the polymerase module in the C-terminal part of the RdRP protein may be regulated in cis by the N-terminal region of the same polypeptide. Either being a methyltransferase in Flavivirus or a functionally unclarified module in Pestivirus, this region could play auxiliary roles for the canonical folding and/or the catalysis of the polymerase, through defined intra-molecular interactions.

No MeSH data available.


Related in: MedlinePlus

Catalytic motifs of Flaviviridae RdRPs. (a) Stereo-pair images of spatial organization of the JEV RdRP catalytic motifs A–G (pdb entry: 4K6M). Seven RdRP motifs are shown as thick noodles. The color-coding is as in Figure 1b; (b) A structure-based sequence alignment depicting the conservation of RdRP motifs (pdb entries: 4K6M, 1S4F, 1NB4, 3OL6, 3BSO, 3AVT [36], 1HI0, 2PGG [37], 4WRT [38], 1RTD [39], and 3DU6 [40]). Three RdRPs from other positive-strand RNA viruses (PICO/PV: Picornaviridae/poliovirus; CALI/NV: Caliciviridae/norovirus; LEVI/QB: Leviviridae/bacteriophage Qβ), two from double-stranded RNA viruses (CYST/PHI6: Cystoviridae/bacteriophage ϕ6; BIRN/BIRV: Birnaviridae/birnavirus), one from negative-stranded RNA viruses (ORTH/IBV: Orthomyxoviridae/influenza virus B), and two reverse transcriptases (RTs) (RETR/HIV1: Retroviridae/human immunodeficiency virus 1; TERT: Telomerase RT) were chosen as representatives for alignment and/or comparison. Only the structurally conserved segment of motif D is included in this alignment. Some important RdRP consensus residues are highlighted in red. Two motif G residues interacting with the +1/+2 junction of the RNA template are indicated by a red box. The two universal aspartic acid residues are indicated by asterisks. Numbers in parenthesis indicate the number of residues not shown. The alignment of IBV polymerase motif G is of lower confidence due to a lower level of structural similarity to motif G in other structures.
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ijms-16-12943-f002: Catalytic motifs of Flaviviridae RdRPs. (a) Stereo-pair images of spatial organization of the JEV RdRP catalytic motifs A–G (pdb entry: 4K6M). Seven RdRP motifs are shown as thick noodles. The color-coding is as in Figure 1b; (b) A structure-based sequence alignment depicting the conservation of RdRP motifs (pdb entries: 4K6M, 1S4F, 1NB4, 3OL6, 3BSO, 3AVT [36], 1HI0, 2PGG [37], 4WRT [38], 1RTD [39], and 3DU6 [40]). Three RdRPs from other positive-strand RNA viruses (PICO/PV: Picornaviridae/poliovirus; CALI/NV: Caliciviridae/norovirus; LEVI/QB: Leviviridae/bacteriophage Qβ), two from double-stranded RNA viruses (CYST/PHI6: Cystoviridae/bacteriophage ϕ6; BIRN/BIRV: Birnaviridae/birnavirus), one from negative-stranded RNA viruses (ORTH/IBV: Orthomyxoviridae/influenza virus B), and two reverse transcriptases (RTs) (RETR/HIV1: Retroviridae/human immunodeficiency virus 1; TERT: Telomerase RT) were chosen as representatives for alignment and/or comparison. Only the structurally conserved segment of motif D is included in this alignment. Some important RdRP consensus residues are highlighted in red. Two motif G residues interacting with the +1/+2 junction of the RNA template are indicated by a red box. The two universal aspartic acid residues are indicated by asterisks. Numbers in parenthesis indicate the number of residues not shown. The alignment of IBV polymerase motif G is of lower confidence due to a lower level of structural similarity to motif G in other structures.

Mentions: For all viral RdRPs, the palm domain harbors polymerase motifs A–E with the most conserved feature embedded in motifs A–C [30,31]. Motifs A and C each contains an aspartic acid residue that is universally conserved for all single-subunit processive nucleic acid polymerases (Figure 2b), playing central roles in the two-metal ion catalytic mechanism [17,32]. Motif B includes a highly conserved serine (JEV NS5 residue 604 or equivalent) known to play key roles in recognizing the 2ʹ-hydroxyl group of the NTP ribose [26,33]. It also contains a glycine residue (JEV NS5 residue 605 or equivalent) that is conserved in all RNA-dependent polymerases. This glycine not only provides flexibility for the Ser-Gly peptide bond flip to coordinate the side-chain rotamer change of the adjacent serine in active site closure [26], but also may play a key role in polymerase translocation. In a recent study, a loop region (JEV NS5 residues 603–609 or equivalent) of PV RdRP centering around this glycine was shown to adopt two distinct backbone conformations and was proposed to mediate the movement of the template-product duplex toward the upstream in the post-catalysis translocation event [34]. Motifs D and E are less conserved in sequence. Structurally, motif D is associated with motif A and they undergo coordinated conformational changes during the closure and reopening of the active site [26]. A motif D lysine residue (K359 in PV RdRP) that is conserved in primer-dependent RdRPs has been proposed to play critical roles in catalysis [35]. However, the region spanning this residue does not have equivalents in de novo RdRPs, either by sequence or by structural homology. Motif E folds with motif C, and primarily interacts with the backbone of the −2 and −3 positions (i.e., the 2nd and the 3rd nucleotide upstream of the active site) of the product RNA.


A Structural Overview of RNA-Dependent RNA Polymerases from the Flaviviridae Family.

Wu J, Liu W, Gong P - Int J Mol Sci (2015)

Catalytic motifs of Flaviviridae RdRPs. (a) Stereo-pair images of spatial organization of the JEV RdRP catalytic motifs A–G (pdb entry: 4K6M). Seven RdRP motifs are shown as thick noodles. The color-coding is as in Figure 1b; (b) A structure-based sequence alignment depicting the conservation of RdRP motifs (pdb entries: 4K6M, 1S4F, 1NB4, 3OL6, 3BSO, 3AVT [36], 1HI0, 2PGG [37], 4WRT [38], 1RTD [39], and 3DU6 [40]). Three RdRPs from other positive-strand RNA viruses (PICO/PV: Picornaviridae/poliovirus; CALI/NV: Caliciviridae/norovirus; LEVI/QB: Leviviridae/bacteriophage Qβ), two from double-stranded RNA viruses (CYST/PHI6: Cystoviridae/bacteriophage ϕ6; BIRN/BIRV: Birnaviridae/birnavirus), one from negative-stranded RNA viruses (ORTH/IBV: Orthomyxoviridae/influenza virus B), and two reverse transcriptases (RTs) (RETR/HIV1: Retroviridae/human immunodeficiency virus 1; TERT: Telomerase RT) were chosen as representatives for alignment and/or comparison. Only the structurally conserved segment of motif D is included in this alignment. Some important RdRP consensus residues are highlighted in red. Two motif G residues interacting with the +1/+2 junction of the RNA template are indicated by a red box. The two universal aspartic acid residues are indicated by asterisks. Numbers in parenthesis indicate the number of residues not shown. The alignment of IBV polymerase motif G is of lower confidence due to a lower level of structural similarity to motif G in other structures.
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Related In: Results  -  Collection

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ijms-16-12943-f002: Catalytic motifs of Flaviviridae RdRPs. (a) Stereo-pair images of spatial organization of the JEV RdRP catalytic motifs A–G (pdb entry: 4K6M). Seven RdRP motifs are shown as thick noodles. The color-coding is as in Figure 1b; (b) A structure-based sequence alignment depicting the conservation of RdRP motifs (pdb entries: 4K6M, 1S4F, 1NB4, 3OL6, 3BSO, 3AVT [36], 1HI0, 2PGG [37], 4WRT [38], 1RTD [39], and 3DU6 [40]). Three RdRPs from other positive-strand RNA viruses (PICO/PV: Picornaviridae/poliovirus; CALI/NV: Caliciviridae/norovirus; LEVI/QB: Leviviridae/bacteriophage Qβ), two from double-stranded RNA viruses (CYST/PHI6: Cystoviridae/bacteriophage ϕ6; BIRN/BIRV: Birnaviridae/birnavirus), one from negative-stranded RNA viruses (ORTH/IBV: Orthomyxoviridae/influenza virus B), and two reverse transcriptases (RTs) (RETR/HIV1: Retroviridae/human immunodeficiency virus 1; TERT: Telomerase RT) were chosen as representatives for alignment and/or comparison. Only the structurally conserved segment of motif D is included in this alignment. Some important RdRP consensus residues are highlighted in red. Two motif G residues interacting with the +1/+2 junction of the RNA template are indicated by a red box. The two universal aspartic acid residues are indicated by asterisks. Numbers in parenthesis indicate the number of residues not shown. The alignment of IBV polymerase motif G is of lower confidence due to a lower level of structural similarity to motif G in other structures.
Mentions: For all viral RdRPs, the palm domain harbors polymerase motifs A–E with the most conserved feature embedded in motifs A–C [30,31]. Motifs A and C each contains an aspartic acid residue that is universally conserved for all single-subunit processive nucleic acid polymerases (Figure 2b), playing central roles in the two-metal ion catalytic mechanism [17,32]. Motif B includes a highly conserved serine (JEV NS5 residue 604 or equivalent) known to play key roles in recognizing the 2ʹ-hydroxyl group of the NTP ribose [26,33]. It also contains a glycine residue (JEV NS5 residue 605 or equivalent) that is conserved in all RNA-dependent polymerases. This glycine not only provides flexibility for the Ser-Gly peptide bond flip to coordinate the side-chain rotamer change of the adjacent serine in active site closure [26], but also may play a key role in polymerase translocation. In a recent study, a loop region (JEV NS5 residues 603–609 or equivalent) of PV RdRP centering around this glycine was shown to adopt two distinct backbone conformations and was proposed to mediate the movement of the template-product duplex toward the upstream in the post-catalysis translocation event [34]. Motifs D and E are less conserved in sequence. Structurally, motif D is associated with motif A and they undergo coordinated conformational changes during the closure and reopening of the active site [26]. A motif D lysine residue (K359 in PV RdRP) that is conserved in primer-dependent RdRPs has been proposed to play critical roles in catalysis [35]. However, the region spanning this residue does not have equivalents in de novo RdRPs, either by sequence or by structural homology. Motif E folds with motif C, and primarily interacts with the backbone of the −2 and −3 positions (i.e., the 2nd and the 3rd nucleotide upstream of the active site) of the product RNA.

Bottom Line: Upon the transition to the elongation phase, this priming element needs to undergo currently unresolved conformational rearrangements to accommodate the growth of the template-product RNA duplex.In the genera of Flavivirus and Pestivirus, the polymerase module in the C-terminal part of the RdRP protein may be regulated in cis by the N-terminal region of the same polypeptide.Either being a methyltransferase in Flavivirus or a functionally unclarified module in Pestivirus, this region could play auxiliary roles for the canonical folding and/or the catalysis of the polymerase, through defined intra-molecular interactions.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Chinese Academy of Sciences, No. 44 Xiao Hong Shan, Wuchang District, Wuhan 430071, China. wujiqin2010@163.com.

ABSTRACT
RNA-dependent RNA polymerases (RdRPs) from the Flaviviridae family are representatives of viral polymerases that carry out RNA synthesis through a de novo initiation mechanism. They share a ≈ 600-residue polymerase core that displays a canonical viral RdRP architecture resembling an encircled right hand with palm, fingers, and thumb domains surrounding the active site. Polymerase catalytic motifs A-E in the palm and motifs F/G in the fingers are shared by all viral RdRPs with sequence and/or structural conservations regardless of the mechanism of initiation. Different from RdRPs carrying out primer-dependent initiation, Flaviviridae and other de novo RdRPs utilize a priming element often integrated in the thumb domain to facilitate primer-independent initiation. Upon the transition to the elongation phase, this priming element needs to undergo currently unresolved conformational rearrangements to accommodate the growth of the template-product RNA duplex. In the genera of Flavivirus and Pestivirus, the polymerase module in the C-terminal part of the RdRP protein may be regulated in cis by the N-terminal region of the same polypeptide. Either being a methyltransferase in Flavivirus or a functionally unclarified module in Pestivirus, this region could play auxiliary roles for the canonical folding and/or the catalysis of the polymerase, through defined intra-molecular interactions.

No MeSH data available.


Related in: MedlinePlus