TY  - JOUR
A1  - Liaqat, Anam
A1  - Stiller, Carina
A1  - Michel, Manuela
A1  - Sednev, Maksim V.
A1  - Höbartner, Claudia
T1  - N\(^6\)-Isopentenyladenosine in RNA Determines the Cleavage Site of Endonuclease Deoxyribozymes
JF  - Angewandte Chemie International Edition
N2  - RNA-cleaving deoxyribozymes can serve as selective sensors and catalysts to examine the modification state of RNA. However, site-specific endonuclease deoxyribozymes that selectively cleave posttranscriptionally modified RNA are extremely rare and their specificity over unmodified RNA is low. In this study, we report that the native tRNA modification N\(^6\)-isopentenyladenosine (i\(^6\)A) strongly enhances the specificity and has the power to reconfigure the active site of an RNA-cleaving deoxyribozyme. Using in vitro selection, we identified a DNA enzyme that cleaves i\(^6\)A-modified RNA at least 2500-fold faster than unmodified RNA. Another deoxyribozyme shows unique and unprecedented behaviour by shifting its cleavage site in the presence of the i\(^6\)A RNA modification. Together with deoxyribozymes that are strongly inhibited by i\(^6\)A, these results highlight intricate ways of modulating the catalytic activity of DNA by posttranscriptional RNA modifications.
KW  - Deoxyribozymes
KW  - Epitranscriptomics
KW  - in vitro selection
KW  - RNA modification
KW  - site-specific RNA cleavage
Y1  - 2020
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-212121
ET  - Early View
ER  - 
TY  - JOUR
A1  - Liaqat, Anam
A1  - Sednev, Maksim V.
A1  - Stiller, Carina
A1  - Höbartner, Claudia
T1  - RNA-cleaving deoxyribozymes differentiate methylated cytidine isomers in RNA
JF  - Angewandte Chemie International Edition
N2  - Deoxyribozymes are emerging as modification-specific endonucleases for the analysis of epigenetic RNA modifications. Here, we report RNA-cleaving deoxyribozymes that differentially respond to the presence of natural methylated cytidines, 3-methylcytidine (m\(^3\)C), N\(^4\)-methylcytidine (m\(^4\)C), and 5-methylcytidine (m\(^5\)C), respectively. Using in vitro selection, we found several DNA catalysts, which are selectively activated by only one of the three cytidine isomers, and display 10- to 30-fold accelerated cleavage of their target m\(^3\)C-, m\(^4\)C- or m\(^5\)C-modified RNA. An additional deoxyribozyme is strongly inhibited by any of the three methylcytidines, but effectively cleaves unmodified RNA. The mXC-detecting deoxyribozymes are programmable for the interrogation of natural RNAs of interest, as demonstrated for human mitochondrial tRNAs containing known m\(^3\)C and m\(^5\)C sites. The results underline the potential of synthetic functional DNA to shape highly selective active sites.
KW  - organic chemistry
KW  - site-specific RNA cleavage
KW  - deoxyribozymes
KW  - epitranscriptomics
KW  - in vitro selection
KW  - RNA modification
Y1  - 2021
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-256519
VL  - 60
ER  - 
TY  - CHAP
A1  - Liaqat, Anam
A1  - Sednev, Maksim V.
A1  - Höbartner, Claudia
T1  - In Vitro Selection of Deoxyribozymes for the Detection of RNA Modifications
T2  - Ribosome Biogenesis: Methods and Protocols
N2  - Deoxyribozymes are artificially evolved DNA molecules with catalytic abilities. RNA-cleaving deoxyribozymes have been recognized as an efficient tool for detection of modifications in target RNAs and provide an alternative to traditional and modern methods for detection of ribose or nucleobase methylation. However, there are only few examples of DNA enzymes that specifically reveal the presence of a certain type of modification, including N6-methyladenosine, and the knowledge about how DNA enzymes recognize modified RNAs is still extremely limited. Therefore, DNA enzymes cannot be easily engineered for the analysis of desired RNA modifications, but are instead identified by in vitro selection from random DNA libraries using synthetic modified RNA substrates. This protocol describes a general in vitro selection stagtegy to evolve new RNA-cleaving DNA enzymes that can efficiently differentiate modified RNA substrates from their unmodified counterpart.
KW  - RNA
KW  - deoxyribozymes
KW  - modified RNA nucleotides
KW  - catalytic DNA
KW  - epitranscriptomics
KW  - in vitro selection
KW  - RNA cleavage
Y1  - 2022
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-279208
SN  - 978-1-0716-2501-9
PB  - Humana Press
ER  -