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Core flexibility of a truncated metazoan mitochondrial tRNA.

Frazer-Abel AA, Hagerman PJ - Nucleic Acids Res. (2008)

Bottom Line: Thus, the absence of canonical TpsiC-D interactions likely results in greater dispersion of anticodon-acceptor interstem angle than for canonical tRNAs.To test this hypothesis, we have assessed the dispersion of the anticodon-acceptor angle for bovine mtRNA(Ser)(AGY), which lacks the canonical D arm and is thus incapable of forming stabilizing interarm interactions.These results suggest that increased flexibility, in addition to a more open interstem angle, would allow both noncanonical and canonical mtRNAs to utilize the same protein synthetic apparatus.

View Article: PubMed Central - PubMed

Affiliation: National Jewish Health, Denver, CO 80206, USA.

ABSTRACT
Secondary and tertiary structures of tRNAs are remarkably preserved from bacteria to humans, the notable exception being the mitochondrial (m) tRNAs of metazoans, which often deviate substantially from the canonical cloverleaf (secondary) or 'L'-shaped (tertiary) structure. Many metazoan mtRNAs lack either the TpsiC (T) or dihydrouridine (D) loops of the canonical cloverleaf, which are known to confer structural rigidity to the folded structure. Thus, the absence of canonical TpsiC-D interactions likely results in greater dispersion of anticodon-acceptor interstem angle than for canonical tRNAs. To test this hypothesis, we have assessed the dispersion of the anticodon-acceptor angle for bovine mtRNA(Ser)(AGY), which lacks the canonical D arm and is thus incapable of forming stabilizing interarm interactions. Using the method of transient electric birefringence (TEB), and by changing the helical torsion angle between a core mtRNA bend and a second bend of known angle/rigidity, we have demonstrated that the core of mtRNA(Ser)(AGY) has substantially greater flexibility than its well-characterized canonical counterpart, yeast cytoplasmic tRNA(Phe). These results suggest that increased flexibility, in addition to a more open interstem angle, would allow both noncanonical and canonical mtRNAs to utilize the same protein synthetic apparatus.

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Relative gel electrophoretic mobility. The outside lanes in all gels contain linear duplex controls of the indicated base pairs in length. Numbering of the intervening lanes corresponds to the number of phasing bases added. Gel conditions described in Materials and methods section.
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Figure 5: Relative gel electrophoretic mobility. The outside lanes in all gels contain linear duplex controls of the indicated base pairs in length. Numbering of the intervening lanes corresponds to the number of phasing bases added. Gel conditions described in Materials and methods section.

Mentions: Heteroduplex RNA constructs containing the bovine mtRNASer(AGY) core element and an A5 bulge, separated by varying numbers of base pairs, were run on polyacrylamide gels (Figure 5, right panels). The weak phase dependence of the mobilities of the two-bend constructs stands in stark contrast to the large phase dependence observed for the yeast tRNAPhe–A5 constructs (Figure 5, left panels), for which previous studies demonstrated substantial structural rigidity of the yeast tRNAPhe core (6). The flexibility contributing to the weak phasing must reside in the bovine mtRNASer(AGY) core element, since the A5 bulge is shared for heteroduplex constructs involving both tRNA species. Interestingly, comparison also reveals that while the canonical yeast tRNAPhe constructs exhibit more pronounced phasing in the presence of magnesium than in its absence, the reverse is true for the noncanonical bovine constructs. This latter observation indicates that magnesium is exerting fundamentally different stabilizing effects for the two tRNAs. It should be noted that these effects of magnesium are only revealed with the current two-bend constructs, where phasing is possible. Determination of the exact nature of the magnesium effects is beyond the scope of this work, and additional studies would be necessary to begin to determine whether the structural modulation is specific to magnesium, or is a more general effect of divalent ions.Figure 5.


Core flexibility of a truncated metazoan mitochondrial tRNA.

Frazer-Abel AA, Hagerman PJ - Nucleic Acids Res. (2008)

Relative gel electrophoretic mobility. The outside lanes in all gels contain linear duplex controls of the indicated base pairs in length. Numbering of the intervening lanes corresponds to the number of phasing bases added. Gel conditions described in Materials and methods section.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2553581&req=5

Figure 5: Relative gel electrophoretic mobility. The outside lanes in all gels contain linear duplex controls of the indicated base pairs in length. Numbering of the intervening lanes corresponds to the number of phasing bases added. Gel conditions described in Materials and methods section.
Mentions: Heteroduplex RNA constructs containing the bovine mtRNASer(AGY) core element and an A5 bulge, separated by varying numbers of base pairs, were run on polyacrylamide gels (Figure 5, right panels). The weak phase dependence of the mobilities of the two-bend constructs stands in stark contrast to the large phase dependence observed for the yeast tRNAPhe–A5 constructs (Figure 5, left panels), for which previous studies demonstrated substantial structural rigidity of the yeast tRNAPhe core (6). The flexibility contributing to the weak phasing must reside in the bovine mtRNASer(AGY) core element, since the A5 bulge is shared for heteroduplex constructs involving both tRNA species. Interestingly, comparison also reveals that while the canonical yeast tRNAPhe constructs exhibit more pronounced phasing in the presence of magnesium than in its absence, the reverse is true for the noncanonical bovine constructs. This latter observation indicates that magnesium is exerting fundamentally different stabilizing effects for the two tRNAs. It should be noted that these effects of magnesium are only revealed with the current two-bend constructs, where phasing is possible. Determination of the exact nature of the magnesium effects is beyond the scope of this work, and additional studies would be necessary to begin to determine whether the structural modulation is specific to magnesium, or is a more general effect of divalent ions.Figure 5.

Bottom Line: Thus, the absence of canonical TpsiC-D interactions likely results in greater dispersion of anticodon-acceptor interstem angle than for canonical tRNAs.To test this hypothesis, we have assessed the dispersion of the anticodon-acceptor angle for bovine mtRNA(Ser)(AGY), which lacks the canonical D arm and is thus incapable of forming stabilizing interarm interactions.These results suggest that increased flexibility, in addition to a more open interstem angle, would allow both noncanonical and canonical mtRNAs to utilize the same protein synthetic apparatus.

View Article: PubMed Central - PubMed

Affiliation: National Jewish Health, Denver, CO 80206, USA.

ABSTRACT
Secondary and tertiary structures of tRNAs are remarkably preserved from bacteria to humans, the notable exception being the mitochondrial (m) tRNAs of metazoans, which often deviate substantially from the canonical cloverleaf (secondary) or 'L'-shaped (tertiary) structure. Many metazoan mtRNAs lack either the TpsiC (T) or dihydrouridine (D) loops of the canonical cloverleaf, which are known to confer structural rigidity to the folded structure. Thus, the absence of canonical TpsiC-D interactions likely results in greater dispersion of anticodon-acceptor interstem angle than for canonical tRNAs. To test this hypothesis, we have assessed the dispersion of the anticodon-acceptor angle for bovine mtRNA(Ser)(AGY), which lacks the canonical D arm and is thus incapable of forming stabilizing interarm interactions. Using the method of transient electric birefringence (TEB), and by changing the helical torsion angle between a core mtRNA bend and a second bend of known angle/rigidity, we have demonstrated that the core of mtRNA(Ser)(AGY) has substantially greater flexibility than its well-characterized canonical counterpart, yeast cytoplasmic tRNA(Phe). These results suggest that increased flexibility, in addition to a more open interstem angle, would allow both noncanonical and canonical mtRNAs to utilize the same protein synthetic apparatus.

Show MeSH
Related in: MedlinePlus