Merck
  • Home
  • Search Results
  • A magnesium-induced RNA conformational switch at the internal ribosome entry site of hepatitis C virus genome visualized by atomic force microscopy.

A magnesium-induced RNA conformational switch at the internal ribosome entry site of hepatitis C virus genome visualized by atomic force microscopy.

Nucleic acids research (2014-12-17)
Ana García-Sacristán, Miguel Moreno, Ascensión Ariza-Mateos, Elena López-Camacho, Rosa M Jáudenes, Luis Vázquez, Jordi Gómez, José Ángel Martín-Gago, Carlos Briones
ABSTRACT

The 5' untranslated region of hepatitis C virus (HCV) genomic RNA contains an internal ribosome entry site (IRES) element, composed of domains II-IV, which is required for cap-independent translation initiation. Little information on the 3D structure of the whole functional HCV IRES is still available. Here, we use atomic force microscopy to visualize the HCV IRES conformation in its natural sequence context, which includes the upstream domain I and the essential, downstream domains V and VI. The 574 nt-long molecule analyzed underwent an unexpected, Mg(2+)-induced switch between two alternative conformations: from 'open', elongated morphologies at 0-2 mM Mg(2+) concentration to a 'closed', comma-shaped conformation at 4-6 mM Mg(2+). This sharp transition, confirmed by gel-shift analysis and partial RNase T1 cleavage, was hindered by the microRNA miR-122. The comma-shaped IRES-574 molecules visualized at 4-6 mM Mg(2+) in the absence of miR-122 showed two arms. Our data support that the first arm would contain domain III, while the second one would be composed of domains (I-II)+(V-VI) thanks to a long-range RNA interaction between the I-II spacer and the basal region of domain VI. This reinforces the previously described structural continuity between the HCV IRES and its flanking domains I, V and VI.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Acrylamide solution, 40%, suitable for electrophoresis, sterile-filtered
Sigma-Aldrich
2-Propanol, BioReagent, for molecular biology, ≥99.5%
Sigma-Aldrich
(3-Aminopropyl)triethoxysilane, ≥98%
Sigma-Aldrich
Glycerol, tested according to Ph. Eur., anhydrous
Supelco
Acrylamide solution, 40% in H2O, for molecular biology
Sigma-Aldrich
2-Propanol, BioUltra, for molecular biology, ≥99.5% (GC)
Supelco
2-Propanol, analytical standard
Supelco
o-Xylene, analytical standard
Sigma-Aldrich
(3-Aminopropyl)triethoxysilane, packaged for use in deposition systems, ≥98%
Sigma-Aldrich
Glycerol, FCC, FG
Sigma-Aldrich
Glycerol, BioReagent, suitable for cell culture, suitable for insect cell culture, suitable for electrophoresis, ≥99% (GC)
Sigma-Aldrich
Glycerol, for molecular biology, ≥99.0%
Sigma-Aldrich
Glycerol, BioXtra, ≥99% (GC)
Sigma-Aldrich
Glycerol, ≥99.5%
Sigma-Aldrich
Glycerol, BioUltra, for molecular biology, anhydrous, ≥99.5% (GC)
Supelco
2-Propanol, Pharmaceutical Secondary Standard; Certified Reference Material
Sigma-Aldrich
(3-Aminopropyl)triethoxysilane, 99%
Sigma-Aldrich
o-Xylene, anhydrous, 97%
Sigma-Aldrich
2-Propanol, anhydrous, 99.5%
Sigma-Aldrich
2-Propanol, electronic grade, 99.999% trace metals basis
Supelco
o-Xylene, Pharmaceutical Secondary Standard; Certified Reference Material
Sigma-Aldrich
(3-Aminopropyl)triethoxysilane, ≥98.0%
Sigma-Aldrich
2-Propanol, suitable for HPLC, 99.9%
Sigma-Aldrich
2-Propanol, HPLC Plus, for HPLC, GC, and residue analysis, 99.9%
Sigma-Aldrich
2-Propanol, suitable for HPLC, 99.5%
Sigma-Aldrich
Glycerol, ReagentPlus®, ≥99.0% (GC)
Sigma-Aldrich
2-Propanol, puriss. p.a., ACS reagent, ≥99.8% (GC)
Sigma-Aldrich
Glycerol, ACS reagent, ≥99.5%
Sigma-Aldrich
2-Propanol, ACS reagent, ≥99.5%
Supelco
Glycerol, analytical standard