Locked Nucleic Acids: Synthesis and Characterization of LNA‐T Diol

Kierzek

et al. 2007

Nucleic Acids Research

Modified nucleotides are useful tools to study the structures, biological functions and chemical and thermodynamic stabilities of nucleic acids. Derivatives of 2,6-diaminopurine riboside (D) are one type of modified nucleotide. The presence of an additional amino group at position 2 relative to adenine results in formation of a third hydrogen bond when interacting with uridine. New method for chemical synthesis of protected 3′-O-phosphoramidite of LNA-2,6-diaminopurine riboside is described. The derivatives of 2′-O-methyl-2,6-diaminopurine and LNA-2,6-diaminopurine ribosides were used to prepare complete 2′-O-methyl RNA and LNA-2′-O-methyl RNA chimeric oligonucleotides to pair with RNA oligonucleotides. Thermodynamic stabilities of these duplexes demonstrated that replacement of a single internal 2′-O-methyladenosine with 2′-O-methyl-2,6-diaminopurine riboside (DM) or LNA-2,6-diaminopurine riboside (DL) increases the thermodynamic stability (ΔΔG°37) on average by 0.9 and 2.3 kcal/mol, respectively. Moreover, the results fit a nearest neighbor model for predicting duplex stability at 37°C. D-A and D-G but not D-C mismatches formed by DM or DL generally destabilize 2′-O-methyl RNA/RNA and LNA-2′-O-methyl RNA/RNA duplexes relative to the same type of mismatches formed by 2′-O-methyladenosine and LNA-adenosine, respectively. The enhanced thermodynamic stability of fully complementary duplexes and decreased thermodynamic stability of some mismatched duplexes are useful for many RNA studies, including those involving microarrays.

Section: Resultsmentioning

confidence: 99%

A chemical synthesis of LNA-2,6-diaminopurine riboside, and the influence of 2′-O-methyl-2,6-diaminopurine and LNA-2,6-diaminopurine ribosides on the thermodynamic properties of 2′-O-methyl RNA/RNA heteroduplexes

Kierzek

et al. 2007

Nucleic Acids Research

“…Synthesis of LNA-U Az phosphoramidite and ONs modified with LNA-U Az LNA-U Az phosphoramidite was prepared from 5-iodo-LNA-U nucleoside derivative 1 synthesized according to the known protocols (Scheme 1). 45,46 Azobenzene was introduced at the C5 position of uracil via an alkyne linker by the Sonogashira coupling reaction between 1 34 and 4-ethynylazobenzene. 47 Phosphitylation of the 3′-hydroxyl group of compound 2 provided the desired phosphoramidite 3.…”

Section: Resultsmentioning

confidence: 99%

C5-azobenzene-functionalized locked nucleic acid uridine: isomerization properties, hybridization ability, and enzymatic stability

et al. 2015

Oligonucleotides (ONs) modified with a locked nucleic acid (LNA) are widely used in the fields of therapeutics, diagnosis, and nanotechnology. There have been significant efforts towards developing LNA analogues bearing modified bridges to improve their hybridization ability, nuclease resistance, and pharmacokinetic profiles. Moreover, nucleobase modifications of LNA are useful strategies for the functionalization of ONs. Modifications of the C5-position of pyrimidine nucleobases are particularly interesting because they enable predictable positioning of functional groups in the major groove of the duplex. Here we report the synthesis of C5-azobenzene-functionalized LNA uridine (LNA-U(Az)) and properties of LNA-U(Az)-modified ONs, including isomerization properties, hybridization ability, and enzyme stability. LNA-U(Az) in ON is photo-isomerized effectively and reversibly by irradiation at 365 nm (trans to cis) and 450 nm (cis to trans). LNA-U(Az)-modified ONs show RNA-selective hybridization ability despite the large hydrophobic azobenzene moiety extending into the major groove of the duplex. The enzymatic stability of LNA-U(Az)-modified ONs is higher than that of natural and LNA-modified ONs with or without photo-irradiation. Our results indicate that LNA-U(Az) holds promise for RNA targeting and photo-switchable technologies.

“…Silica gel column purified N-glycoside only was used for the next reaction. The next reactions were performed according to the original procedure, except: in exchanging of 5'-O-methanosulfonyl, lithium benzoate was substituted for sodium benzoate due to solubility in DMF (12, 17, 30). A two-step deprotection was used for removal of 3'-O-allyl (44).…”

Section: Resultsmentioning

confidence: 99%

Chemical Synthesis of LNA-2-thiouridine and Its Influence on Stability and Selectivity of Oligonucleotide Binding to RNA

et al. 2009

Hybridization to RNA is important for many applications, including antisense therapeutics, RNA interference, and microarray screening. Similar thermodynamic stabilities of A-U and G-U base pairs result in difficulties in selective binding of matched and mismatched RNA duplexes. Moreover, A-U pairs are weaker than G-C pairs so that binding is sometimes weak when many A-U pairs are present. It is known, however, that replacement of uridine with 2-thiouridine significantly improves binding and selectivity. To test for additional improvement of binding and of the specificity for binding A over G, LNA-2-thiouridine was synthesized for the first time and incorporated into many LNA-2-O-methylRNA/RNA duplexes. UV melting was used to measure the thermodynamic effect of replacing 2'-O-methyluridine with 2'-O-methyl-2-thiouridine or LNA-2-thiouridine. The 2-thiouridine usually enhances binding and selectivity. Selectivity is optimized when a single 2-thiouridine is placed at an internal position in a duplex.