Limits...
Toward understanding the conformational dynamics of RNA ligation.

Swift RV, Durrant J, Amaro RE, McCammon JA - Biochemistry (2009)

Bottom Line: This study describes a recent 70 ns molecular dynamics simulation of TbREL1, an ATP-dependent RNA-editing ligase of the nucleotidyltransferase superfamily that is required for the survival of T. brucei insect and bloodstream forms.In this work, a model of TbREL1 in complex with its full double-stranded RNA (dsRNA) substrate is created on the basis of the homologous relation between TbREL1 and T4 Rnl2.Important features of RNA binding and specificity are revealed for kinetoplastid ligases and the broader nucleotidyltransferase superfamily.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, NSF Center for Theoretical Biological Physics, University of California at San Diego, La Jolla, California 92093-0365, USA.

ABSTRACT
Members of the genus Trypanosoma, which include the pathogenic species Trypanosoma brucei and Trypanosoma cruzi, edit their post-transcriptional mitochondrial RNA via a multiprotein complex called the editosome. In T. brucei, the RNA is nicked prior to uridylate insertion and deletion. Following editing, nicked RNA is religated by one of two RNA-editing ligases (TbREL). This study describes a recent 70 ns molecular dynamics simulation of TbREL1, an ATP-dependent RNA-editing ligase of the nucleotidyltransferase superfamily that is required for the survival of T. brucei insect and bloodstream forms. In this work, a model of TbREL1 in complex with its full double-stranded RNA (dsRNA) substrate is created on the basis of the homologous relation between TbREL1 and T4 Rnl2. The simulation captures TbREL1 dynamics in the state immediately preceding RNA ligation, providing insights into the functional dynamics and catalytic mechanism of the kinetoplastid ligation reaction. Important features of RNA binding and specificity are revealed for kinetoplastid ligases and the broader nucleotidyltransferase superfamily.

Show MeSH
RNA binding footprint. In the top panel, the protein dynamic contact footprint is projected onto the molecular surface of the RNA. In the bottom panel, the RNA dynamic contact footprint is projected onto the molecular surface of TbREL1. Both the protein and RNA are shown in gray surface representation with the contacts colored by percent occupancy over the dynamics portion of the simulation. A unique kinetoplastid insert is outlined in black, and a cluster of residues thought to be important in RNA recognition on the 5′-PO4 side of the nick is outlined in red.
© Copyright Policy - open-access - ccc-price
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2651658&req=5

fig5: RNA binding footprint. In the top panel, the protein dynamic contact footprint is projected onto the molecular surface of the RNA. In the bottom panel, the RNA dynamic contact footprint is projected onto the molecular surface of TbREL1. Both the protein and RNA are shown in gray surface representation with the contacts colored by percent occupancy over the dynamics portion of the simulation. A unique kinetoplastid insert is outlined in black, and a cluster of residues thought to be important in RNA recognition on the 5′-PO4 side of the nick is outlined in red.

Mentions: To identify molecular specificity determinants that mediate dsRNA binding, we generated a RNA binding footprint based on a set of protein−RNA interaction persistence times (Figure 5). This analysis allows us to identify primary contact sites between the RNA and the protein and to discriminate longer-lived interactions from more transient ones. The footprint-like mapping of the protein−RNA interaction persistence times onto the protein and RNA surfaces shows that more extensive interactions occur on the 5′-PO4 side of the nick. Although this contact pattern contrasts with the results of Nandakumar and co-workers, who report that the closest homologue of TbREL1, T4 Rnl2, makes more extensive contacts with the strands on the 3′-OH side of the nick, it supports the experimental observations that TbREL1 requires RNA on the 5′-PO4 strand of the nick for in vitro polynucleotide religation. A large, contiguous set of interactions, as well as various individual interactions, makes up the more extensive contacts on the 5′-PO4 side of the nick. Specifically, D62, L63, P64, S65, S67, and Q70 collectively form a contiguous surface, herein called the “recognition sequence”, which intercalates the minor grove of the dsRNA on the 5′-PO4 side of the nick (Figures 1 and 5). Although the recognition sequence is not conserved at the superfamily level, it is conserved among the Trypanosomatidae type I mitochondrial RNA-editing ligase sequences available in the SWISS-PROT database (39). As the depth and width of the minor grooves differ between the canonical RNA and DNA forms, this recognition sequence may play a key role in polynucleotide discrimination.


Toward understanding the conformational dynamics of RNA ligation.

Swift RV, Durrant J, Amaro RE, McCammon JA - Biochemistry (2009)

RNA binding footprint. In the top panel, the protein dynamic contact footprint is projected onto the molecular surface of the RNA. In the bottom panel, the RNA dynamic contact footprint is projected onto the molecular surface of TbREL1. Both the protein and RNA are shown in gray surface representation with the contacts colored by percent occupancy over the dynamics portion of the simulation. A unique kinetoplastid insert is outlined in black, and a cluster of residues thought to be important in RNA recognition on the 5′-PO4 side of the nick is outlined in red.
© Copyright Policy - open-access - ccc-price
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651658&req=5

fig5: RNA binding footprint. In the top panel, the protein dynamic contact footprint is projected onto the molecular surface of the RNA. In the bottom panel, the RNA dynamic contact footprint is projected onto the molecular surface of TbREL1. Both the protein and RNA are shown in gray surface representation with the contacts colored by percent occupancy over the dynamics portion of the simulation. A unique kinetoplastid insert is outlined in black, and a cluster of residues thought to be important in RNA recognition on the 5′-PO4 side of the nick is outlined in red.
Mentions: To identify molecular specificity determinants that mediate dsRNA binding, we generated a RNA binding footprint based on a set of protein−RNA interaction persistence times (Figure 5). This analysis allows us to identify primary contact sites between the RNA and the protein and to discriminate longer-lived interactions from more transient ones. The footprint-like mapping of the protein−RNA interaction persistence times onto the protein and RNA surfaces shows that more extensive interactions occur on the 5′-PO4 side of the nick. Although this contact pattern contrasts with the results of Nandakumar and co-workers, who report that the closest homologue of TbREL1, T4 Rnl2, makes more extensive contacts with the strands on the 3′-OH side of the nick, it supports the experimental observations that TbREL1 requires RNA on the 5′-PO4 strand of the nick for in vitro polynucleotide religation. A large, contiguous set of interactions, as well as various individual interactions, makes up the more extensive contacts on the 5′-PO4 side of the nick. Specifically, D62, L63, P64, S65, S67, and Q70 collectively form a contiguous surface, herein called the “recognition sequence”, which intercalates the minor grove of the dsRNA on the 5′-PO4 side of the nick (Figures 1 and 5). Although the recognition sequence is not conserved at the superfamily level, it is conserved among the Trypanosomatidae type I mitochondrial RNA-editing ligase sequences available in the SWISS-PROT database (39). As the depth and width of the minor grooves differ between the canonical RNA and DNA forms, this recognition sequence may play a key role in polynucleotide discrimination.

Bottom Line: This study describes a recent 70 ns molecular dynamics simulation of TbREL1, an ATP-dependent RNA-editing ligase of the nucleotidyltransferase superfamily that is required for the survival of T. brucei insect and bloodstream forms.In this work, a model of TbREL1 in complex with its full double-stranded RNA (dsRNA) substrate is created on the basis of the homologous relation between TbREL1 and T4 Rnl2.Important features of RNA binding and specificity are revealed for kinetoplastid ligases and the broader nucleotidyltransferase superfamily.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, NSF Center for Theoretical Biological Physics, University of California at San Diego, La Jolla, California 92093-0365, USA.

ABSTRACT
Members of the genus Trypanosoma, which include the pathogenic species Trypanosoma brucei and Trypanosoma cruzi, edit their post-transcriptional mitochondrial RNA via a multiprotein complex called the editosome. In T. brucei, the RNA is nicked prior to uridylate insertion and deletion. Following editing, nicked RNA is religated by one of two RNA-editing ligases (TbREL). This study describes a recent 70 ns molecular dynamics simulation of TbREL1, an ATP-dependent RNA-editing ligase of the nucleotidyltransferase superfamily that is required for the survival of T. brucei insect and bloodstream forms. In this work, a model of TbREL1 in complex with its full double-stranded RNA (dsRNA) substrate is created on the basis of the homologous relation between TbREL1 and T4 Rnl2. The simulation captures TbREL1 dynamics in the state immediately preceding RNA ligation, providing insights into the functional dynamics and catalytic mechanism of the kinetoplastid ligation reaction. Important features of RNA binding and specificity are revealed for kinetoplastid ligases and the broader nucleotidyltransferase superfamily.

Show MeSH