The conformations of locked nucleic acids (LNA)

Abstract

We have used 2D NMR spectroscopy to study the sugar conformations of oligonucleotides containing a conformationally restricted nucleotide (LNA) with a 2'-O, 4'-C-methylene bridge. We have investigated a modified 9-mer single stranded oligonucleotide as well as three 9- and 10-mer modified oligonucleotides hybridized to unmodified DNA. The single-stranded LNA contained three modifications whereas the duplexes contained one, three and four modifications, respectively. The LNA:DNA duplexes have normal Watson-Crick base-pairing with all the nucleotides in anti-conformation. By use of selective DQF-COSY spectra we determined the ratio between the N-type (C3'-endo) and S-type (C2'-endo) sugar conformations of the nucleotides. In contrast to the corresponding single-stranded DNA (ssDNA), we found that the sugar conformations of the single-stranded LNA oligonucleotide (ssLNA) cannot be described by a major S-type conformer of all the nucleotides. The nucleotides flanking an LNA nucleotide have sugar conformations with a significant population of the N-type conformer. Similarly, the sugar conformations of the nucleotides in the LNA:DNA duplexes flanking a modification were also shown to have significant contributions from the N-type conformation. In all cases, the sugar conformations of the nucleotides in the complementary DNA strand in the duplex remain in the S-type conformation. We found that the locked conformation of the LNA nucleotides both in ssLNA and in the duplexes organize the phosphate backbone in such a way as to introduce higher population of the N-type conformation. These conformational changes are associated with an improved stacking of the nucleobases. Based on the results reported herein, we propose that the exceptional stability of the LNA modified duplexes is caused by a quenching of concerted local backbone motions (preorganization) by the LNA nucleotides in ssLNA so as to decrease the entropy loss on duplex formation combined with a more efficient stacking of the nucleobases.

MicroRNAs: Novel Biomarkers for Human Cancer

“…This problem has been overcome by the use of locked nucleic acids (LNAs). LNAs contain at least one LNA monomer-a nucleic acid analog in which the 2Ј oxygen atom and the 4Ј carbon atom of the ribose moiety is "locked" by a connecting methylene bridge (16 ). Each incorporated LNA monomer increases the T m of the nucleic acid duplex by 2-10°C (17 ).…”

DNAzyme technology and cancer therapy: cleave and let die

“…Locked nucleic acid bases comprise a 2 ¶-O 4-C methylene bridge that locks in a C3 ¶-endo conformation (27), effectively increasing affinity for complementary sequences (28,29). The advantages of locked nucleic acid include increased thermal stability of duplexes toward complementary DNA or RNA, stability toward 3 ¶-exonucleolytic degradation, and greater solubility due to structural similarities to nucleic acids (30,31). Locked nucleic acid incorporation into DNAzymes, however, has been found to influence both catalytic activity under single turnover conditions (24,26) and biological potency (26,32).…”

Interaction of the Ty3 Reverse Transcriptase Thumb Subdomain with Template-Primer

“…Selected residues within this putative ␣-helix of Ty3 RT were mutated and evaluated with respect to the DNA polymerase and RNase H functions of the purified enzymes. In addition to studying duplex DNA and RNA/DNA hybrids, polymerization efficiency on duplex DNA containing targeted insertion of locked nucleic acid analogs (35) or abasic lesions (36,37) …”