Structural studies of trypanosoma brucei RNA editing ligases and their binding partner proteins

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DOIResolve DOI: https://doi.org/10.1021/acs.biochem.5b01257
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Name affiliation
  1. National Research Council of Canada. Human Health Therapeutics
FormatText, Article
Journal titleBiochemistry
ISSN0006-2960
1520-4995
Volume55
Issue16
Pages2319–2331
Abstract
To study the mechanism of ligating nicked RNA strands, we conducted molecular dynamics simulations of Trypanosoma brucei RNA editing ligases L1 and L2 complexed with double-stranded RNA (dsRNA) fragments. In each resulting model, a Mg²⁺ ion coordinates the 5′-PO₄ of the nicked nucleotide and the 3′-OH of the terminal nucleotide for a nucleophilic reaction consistent with the postulated step 3 chemistry of the ligation mechanism. Moreover, coordination of the 3′-OH to the Mg²⁺ ion may lower its pKa, thereby rendering it a more effective nucleophile as an oxyanion. Thus, Mg²⁺ may play a twofold role: bringing the reactants into the proximity of each other and activating the nucleophile. We also conducted solvated interaction energy calculations to explore whether ligation specificities can be correlated to ligase–dsRNA binding affinity changes. The calculated dsRNA binding affinities are stronger for both L1 and L2 when the terminal nucleotide is changed from cytosine to guanine, in line with their experimentally measured ligation specificities. Because the ligation mechanism is also influenced by interactions of the ligase with partner proteins from the editosome subcomplex, we also modeled the structure of the RNA-bound L2 in complex with the oligonucleotide binding (OB) domain of largest editosome interacting protein A1. The resulting L2–dsRNA–A1 model, which is consistent with mutagenesis and binding data recorded to date, provides the first atomic-level glimpse of plausible interactions around the RNA ligation site in the presence of an OB domain presented in-trans to a nucleic acid ligase.
Publication date
PublisherACS Publications
LanguageEnglish
Peer reviewedYes
NRC numberNRC-HHT-53312
NPARC number23000345
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Record created2016-07-08
Record modified2020-03-16
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