Magnesium-binding architectures in RNA crystal structures: validation, binding preferences, classification and motif detection

Zheng, Heping; Shabalin, I.G.; Handing, K.B.; Bujnicki, Janusz; Minor, Władek

doi:10.1093/nar/gkv225

Cited by 96 publications

(213 citation statements)

References 37 publications

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“…The observation that >500 Fe or Mn ions can associate with a bacterial ribosome is consistent with the number of Mg 2+ ions observed by X-ray diffraction (100-1000 Mg 2+ per ribosome (41)), and supports a model in which Fe 2+ or Mn 2+ has replaced Mg 2+ as the dominant divalent cation in our experiments. The high capacity of ribosomes for Fe 2+ and Mn 2+ reflects all rRNA-associated divalent cations, including condensed, glassy and chelated divalent cations (42), and in addition, we presume that Fe 2+ or Mn 2+ can associate with a variety of rProteins, including those previously shown to bind Zn 2+ (e.g.…”

Section: Discussionsupporting

confidence: 89%

Multiple prebiotic metals mediate translation

Bray

Lenz

Haynes

et al. 2018

Preprint

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15In extant biochemistry, Mg 2+ ions are essential for both structure and function of the ribosome (4,5) and for 16functions of many enzymes involved in translation (6). The translation system, which synthesizes all coded 17 protein (7), originated and matured during the Archean Eon (4-2.5 Ga; (8)). The common core of the 18 ribosome and many other aspects of the translation system have remained essentially frozen since the last 19 universal common ancestor (9,10). 20In ribosomes, structural Mg 2+ ions occur in rRNA-Mg 2+ clamps (11) (Fig. 1a), in dinuclear Mg 2+ -21 microclusters that frame the peptidyl transferase center (12) (Fig. 1b), and at the SSU-LSU interface (13) 22( Fig. 1c). Functional Mg 2+ ions stabilize a critical bend in mRNA between the P-site and A-site codons (14) 23 (Fig. 1d), and mediate rRNA-tRNA/mRNA interactions (15) (Fig. 1e, f 26All rights reserved. No reuse allowed without permission.(which was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. (which was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. 97RO). Thus, a total of 2.5 mM "background" Mg 2+ was present in each reaction (Fig. S1) MgCl2 and FeCl2 which were tested over a range of final concentrations (1, 3, 4, 5, 6, 7, 8, 9, and 11 mM; 122 All rights reserved. No reuse allowed without permission.(which was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. the amounts of metals in our assay reaction, which had no effect on DHFR activity (data not shown). 136Ribosome iron content. In order to measure the iron content of ribosomes, a 300 µL reaction was prepared 137 containing 45 µL of a 13.3 µM stock of E. coli ribosomes (New England Biolabs catalog # P0763S), 7 mM

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Section: Discussionsupporting

confidence: 89%

Multiple prebiotic metals mediate translation

Bray

Lenz

Haynes

et al. 2018

Preprint

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“…Mg 2+ has the same number of electrons as water and Na + , which cannot be distinguished from Mg 2+ by difference electron density maps alone. 50 …”

Section: Resultsmentioning

confidence: 99%

Peptide Bond Formation Mechanism Catalyzed by Ribosome

et al. 2015

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In this paper we present a study of the peptide bond formation reaction catalyzed by ribosome. Different mechanistic proposals have been explored by means of Free Energy Perturbation methods within hybrid QM/MM potentials, where the chemical system has been described by the M06-2X functional and the environment by means of the AMBER force field. According to our results, the most favourable mechanism in the ribosome would proceed through an eight-membered ring transition state, involving a proton shuttle mechanism through the hydroxyl group of the sugar and a water molecule. This transition state is similar to that described for the reaction in solution (J. Am. Chem. Soc. 2013, 135, 8708–8719) but the reaction mechanisms are noticeable different. Our simulations reproduce the experimentally determined catalytic effect of ribosome that can be explained by the different behaviour of the two environments. While the solvent reorganizes during the chemical process involving an entropic penalty, the ribosome is preorganized in the formation of the Michaelis complex and does not suffer important changes along the reaction, dampening the charge redistribution of the chemical system.

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“…In addition, in contrast to the deleterious effect of abasic modifications in a small ribozyme, 78 our data demonstrate that in large RNAs the relatively stable RNA abasic 79 modification provides a new strategy to probe functions of conserved nucleobases, analogous to the alanine scanning mutagenesis strategy used to study proteins. 80 Therefore, this work lays the foundation for exploring the thousands of inner-sphere Mg(II) interactions with RNA nucleobases 22 and with the fast-growing list of functional RNA modifications.…”

Section: Discussionmentioning

confidence: 99%

Inner-Sphere Coordination of Divalent Metal Ion with Nucleobase in Catalytic RNA

2017

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Identification of the function of metal ions and the RNA moieties, particularly nucleobases, that bind metal ions are important questions in RNA catalysis. Here we combine single-atom and abasic substitutions to probe functions of conserved nucleobases in ribonuclease P (RNase P). Structural and biophysical studies of bacterial RNase P propose direct coordination of metal ions by the nucleobases of conserved uridine and guanosine in helix P4 of the RNA subunit (P RNA). To biochemically probe the function of metal ion interactions, we substituted the universally conserved bulged uridine (U51) in the P4 helix of circularly permuted Bacillus subtilis P RNA with 4-thiouridine, 4-deoxyuridine and abasic modifications and G378/379 with 2-aminopurine, N7-deazaguanosine, and 6-thioguanosine. The functional group modifications of U51 decrease RNase P-catalyzed phosphodiester bond cleavage 16- to 23-fold, as measured by the single-turnover cleavage rate constant. The activity of the 4-thiouridine RNase P is partially rescued by addition of Cd(II) or Mn(II) ions. This is the first time a metal-rescue experiment provides evidence for inner-sphere divalent metal ion coordination with a nucleobase. Modifications of G379 modestly decrease the cleavage activity of RNase P, suggesting outer-sphere coordination of O6 on G379 to a metal ion. These data provide biochemical evidence for catalytically important interactions of the P4 helix of P RNA with metal ions, demonstrating that the bulged uridine coordinates at least one catalytic metal ion through an inner-sphere interaction. The combination of single-atom and abasic nucleotide substitutions provides a powerful strategy to probe functions of conserved nucleobases in large RNAs.

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Magnesium-binding architectures in RNA crystal structures: validation, binding preferences, classification and motif detection

Cited by 96 publications

References 37 publications

Multiple prebiotic metals mediate translation

Multiple prebiotic metals mediate translation

Peptide Bond Formation Mechanism Catalyzed by Ribosome

Inner-Sphere Coordination of Divalent Metal Ion with Nucleobase in Catalytic RNA

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