TY  - JOUR
A1  - Yong-Boum, Kim
A1  - Wacker, Anna
A1  - Laer, Karl von
A1  - Rogov, Vladimir V.
A1  - Süß, Beatrix
A1  - Schwalbe, Harald
T1  - Ligand binding to 2΄-deoxyguanosine sensing riboswitch in metabolic context
N2  - The mfl-riboswitch is a transcriptional off-switch, which down-regulates expression of subunit ß of ribonucleotide reductase in Mesoplasma florum upon 2´-deoxyguanosine binding. We characterized binding of 2´-deoxyguanosine to the mfl-aptamer domain (WT aptamer) and a sequence-stabilized aptamer (MT aptamer) under in vitro and ‘in-cell-like’ conditions by isothermal titration calorimetry (ITC) and nuclear magnetic resonance (NMR) spectroscopy. ‘In-celllike’ environment was simulated by Bacillus subtilis cell extract, in which both aptamers remained sufficiently stable to detect the resonances of structural elements and ligand binding in 2D NMR experiments. Under ‘in-cell-like’-environment, (i) the WT aptamer bound the endogenous metabolite guanosine and (ii) 2´-deoxyguanosine efficiently displaced guanosine from the WT aptamer. In contrast, MT aptamer exhibited moderate binding to 2´-deoxyguanosine and weak binding to guanosine. NMR experiments indicated that binding of guanosine was not limited to the aptamer domain of the riboswitch but also the full-length mfl-riboswitch bound guanosine, impacting on the regulation efficiency of the riboswitch and hinting that, in addition to 2´-deoxyguanosine, guanosine plays a role in riboswitch function in vivo. Reporter gene assays in B. subtilis demonstrated the regulation capacity of the WT aptamer, whereas the MT aptamer with lower affinity to 2´ -deoxyguanosine was not able to regulate gene expression.
Y1  - 2017
UR  - http://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/43913
UR  - https://nbn-resolving.org/urn:nbn:de:hebis:30:3-439133
N1  - This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited.
VL  - 45
IS  - 9
SP  - 5375
EP  - 5386
PB  - Oxford University Press
CY  - London
ER  -