Locked onto the Target”: Increasing the Efficiency of Antagomirzymes Using Locked Nucleic Acid Modifications
IR@IGIB: CSIR-Institute of Genomics & Integrative Biology, New Delhi
View Archive Info| Field | Value | |
| Title |
Locked onto the Target”: Increasing the Efficiency of Antagomirzymes Using Locked
Nucleic Acid Modifications
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| Creator |
Kaur, Harleen
Scaria, Vinod Maiti, Souvik |
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| Subject |
G1 Genome informatics (General)
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| Description |
This study highlights the effect of incorporation of locked nucleic acid (LNA) on improving the
functional efficacy of DNAzymes against microRNAs (antagomirzymes). DNAzymes were designed against
two different sites of miR-27a, which were encompassed both within the precursor and mature form of
miRNA. The cleavage and functional activities of these DNAzymes have been compared to those of LNA-
modified counterparts, containing LNA modification in each of the substrate binding arms. Preliminary
examination based on in vitro cleavage demonstrated LNAzyme to be much more effective in the ensuing
cleavage of target miRNA under both single- and multiple-turnover conditions. Evaluation of kinetic
parameters indicated almost 5-fold higher cleavage efficiency, kobs, for LNAzymes than for DNAzymes,
leading to more efficient cleavage of the substrate. We attribute this enhancement in cleavage efficiency to the
LNA-mediated improvement in the hybridization of the antagomirzyme 3 target complex. Functional
validation of the relative activities was accomplished through the luciferase reporter assay and quantitative
real-time polymerase chain reaction (qRT-PCR). Both the unmodified and LNA-modified antagomirzymes
were very active in ensuing efficient miRNA knockdown; however, compared to the DNAzymes, the
LNAzymes were almost 25% more active. A direct quantitative estimate of miRNA cleavage, conducted
using qRT-PCR, further substantiated the data by indicating that LNAzyme effectively downregulated the
levels of mature miRNA (up to 50%) versus the corresponding DNAzymes. Our data thus provide formative
evidence of the successful employment of LNA-based antagomirzymes against miRNA.
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| Date |
2010-09-09
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| Type |
Article
PeerReviewed |
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| Format |
application/pdf
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| Identifier |
http://openaccess.igib.res.in/99/1/bi101185k.pdf
Kaur, Harleen and Scaria, Vinod and Maiti, Souvik (2010) Locked onto the Target”: Increasing the Efficiency of Antagomirzymes Using Locked Nucleic Acid Modifications. PubMed, 49 (44). pp. 9449-9456. |
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| Relation |
http://openaccess.igib.res.in/99/
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