Abstract
Thionucleotides, especially 4-thiouridine and 6-thioguanosine, are photosensitive molecules that photocrosslink to both proteins and nucleic acids, and this feature is a major reason for their application in various investigations. To get insight into the thermodynamic and structural contributions of 6-thioguanosine to the properties of RNA duplexes a systematic study was performed. In a series of RNA duplexes, selected guanosine residues located in G-C base pairs, mismatches (G-G, G-U, and G-A), or 5′ and 3′-dangling ends were replaced with 6-thioguanosine. Generally, the presence of 6-thioguanosine diminishes the thermodynamic stability of RNA duplexes. This effect depends on its position within duplexes and the sequence of adjacent base pairs. However, when placed at a dangling end a 6-thioguanosine residue actually exerts a weak stabilizing effect. Furthermore, the structural effect of 6-thioguanosine substitution appears to be minimal based on NMR and Circular Dichroism (CD) data.
Highlights
RNA contains many modified nucleotides performing various biological functions, and thionucleotides constitute one group of these modified nucleotides
Studies of RNA duplex formation by oligonucleotides with either an internal or a terminal 4-thiouridine showed an increase in their thermodynamic stability and indicated that the presence of s4U increases the stability of base pairing with guanosine more than that with
For the same set of oligonucleotides modified with 2-thiouridine (s2U), it has been shown that s2U increases the stability of base pairing with adenosine more than that with guanosine
Summary
The chemical stepwise synthesis of 3′-O-phosphoramidite of the protected 6-thioguanosine is described in detail in Supplementary Materials. Thermodynamic measurements were performed for nine different concentrations of RNA duplex in the range of 10−3–10−6 M on a JASCO V-650 UV/Vis spectrophotometer in buffer containing 1 M sodium chloride, 20 mM sodium cacodylate, and 0.5 mM Na2EDTA (pH 7)[25]. Oligonucleotides were dissolved in a buffer containing 1 M sodium chloride, 20 mM sodium cacodylate and 0.5 mM Na2EDTA (pH 7.0) to achieve a 20 μM sample concentration. For the acquisition of NMR data, all RNA duplexes were dissolved in a 10 mM sodium phosphate buffer (pH 6.8) containing 150 mM sodium chloride and 0.1 mM Na2EDTA.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
