TY  - JOUR
A1  - Mieczkowski, Mateusz
A1  - Steinmetzger, Christian
A1  - Bessi, Irene
A1  - Lenz, Ann-Kathrin
A1  - Schmiedel, Alexander
A1  - Holzapfel, Marco
A1  - Lambert, Christoph
A1  - Pena, Vladimir
A1  - Höbartner, Claudia
T1  - Large Stokes shift fluorescence activation in an RNA aptamer by intermolecular proton transfer to guanine
JF  - Nature Communications
N2  - Fluorogenic RNA aptamers are synthetic functional RNAs that specifically bind and activate conditional fluorophores. The Chili RNA aptamer mimics large Stokes shift fluorescent proteins and exhibits high affinity for 3,5-dimethoxy-4-hydroxybenzylidene imidazolone (DMHBI) derivatives to elicit green or red fluorescence emission. Here, we elucidate the structural and mechanistic basis of fluorescence activation by crystallography and time-resolved optical spectroscopy. Two co-crystal structures of the Chili RNA with positively charged DMHBO+ and DMHBI+ ligands revealed a G-quadruplex and a trans-sugar-sugar edge G:G base pair that immobilize the ligand by π-π stacking. A Watson-Crick G:C base pair in the fluorophore binding site establishes a short hydrogen bond between the N7 of guanine and the phenolic OH of the ligand. Ultrafast excited state proton transfer (ESPT) from the neutral chromophore to the RNA was found with a time constant of 130 fs and revealed the mode of action of the large Stokes shift fluorogenic RNA aptamer.
KW  - Fluorogenic RNA Aptamers
KW  - Synthetic Functional RNAs
KW  - Chili RNA Aptamer
KW  - Co-Crystal Structures of Chili RNA
KW  - RNA
KW  - Optical Spectroscopy
KW  - Structural Biology
KW  - X-ray Crystallography
Y1  - 2021
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-254527
VL  - 12
ER  - 
TY  - JOUR
A1  - Mieczkowski, Mateusz
A1  - Steinmetzger, Christian
A1  - Bessi, Irene
A1  - Lenz, Ann-Kathrin
A1  - Schmiedel, Alexander
A1  - Holzapfel, Marco
A1  - Lambert, Christoph
A1  - Pena, Vladimir
A1  - Höbartner, Claudia
T1  - Large Stokes shift fluorescence activation in an RNA aptamer by intermolecular proton transfer to guanine
JF  - Nature Communications
N2  - Fluorogenic RNA aptamers are synthetic functional RNAs that specifically bind and activate conditional fluorophores. The Chili RNA aptamer mimics large Stokes shift fluorescent proteins and exhibits high affinity for 3,5-dimethoxy-4-hydroxybenzylidene imidazolone (DMHBI) derivatives to elicit green or red fluorescence emission. Here, we elucidate the structural and mechanistic basis of fluorescence activation by crystallography and time-resolved optical spectroscopy. Two co-crystal structures of the Chili RNA with positively charged DMHBO+ and DMHBI+ ligands revealed a G-quadruplex and a trans-sugar-sugar edge G:G base pair that immobilize the ligand by π-π stacking. A Watson-Crick G:C base pair in the fluorophore binding site establishes a short hydrogen bond between the N7 of guanine and the phenolic OH of the ligand. Ultrafast excited state proton transfer (ESPT) from the neutral chromophore to the RNA was found with a time constant of 130 fs and revealed the mode of action of the large Stokes shift fluorogenic RNA aptamer.
KW  - RNA
KW  - optical spectroscopy
KW  - structural biology
KW  - X-ray crystallography
Y1  - 2021
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-270274
VL  - 12
ER  - 
TY  - INPR
A1  - Scheitl, Carolin P. M.
A1  - Mieczkowski, Mateusz
A1  - Schindelin, Hermann
A1  - Höbartner, Claudia
T1  - Structure and mechanism of the methyltransferase ribozyme MTR1
T2  - Nature Chemical Biology
N2  - RNA-catalysed RNA methylation was recently shown to be part of the catalytic repertoire of ribozymes. The methyltransferase ribozyme MTR1 catalyses the site-specific synthesis of 1-methyladenosine (m\(^1\)A) in RNA, using O\(^6\)-methylguanine (m\(^6\)G) as methyl group donor. Here we report the crystal structure of MTR1 at a resolution of 2.8 Å, which reveals a guanine binding site reminiscent of natural guanine riboswitches. The structure represents the postcatalytic state of a split ribozyme in complex with the m1A-containing RNA product and the demethylated cofactor guanine. The structural data suggest the mechanistic involvement of a protonated cytidine in the methyl transfer reaction. A synergistic effect of two 2'-O-methylated ribose residues in the active site results in accelerated methyl group transfer. Supported by these results, it seems plausible that modified nucleotides may have enhanced early RNA catalysis and that metabolite-binding riboswitches may resemble inactivated ribozymes that have lost their catalytic activity during evolution.
KW  - Methyltransferase Ribozyme MTR1
KW  - Crystal structure of MTR1
KW  - RNA-catalyzed RNA methylation
KW  - X-ray crystallography
KW  - RNA
Y1  - 2022
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-272170
ET  - submitted version
ER  -