Faculty of Biochemistry, Chemistry and Pharmacy: Research Data
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- Research DataInvestigating the structural heterogeneity and dynamics in the apical stem loop of s2m from SARS CoV-2 Delta by NMR spectroscopy, MD simulations and SAXS2025-02-28RNAs can fold into heterogeneous structural ensembles populating a complex energy landscape. Structural analysis of RNA loops poses particular challenges and the relationships between their primary sequence, structure, and energy of loops remains enigmatic. We here report the characterisation of the conformational heterogeneity and dynamics of the apical loop in the stem loop 2 motif (s2m) of SARS-CoV-2 (SCoV-2) Delta using a combination of nuclear magnetic resonance spectroscopy (NMR) and molecular dynamics simulations (MD). The apical nonaloop of s2m is flexible and exhibits substantial dynamics, rendering representation of the structure by a single conformation unsuitable. Thus, we report an ensemble of NMR structures in best agreement with SAXS data and weighted MD simulations enabling the representation of the conformational space in the s2m nonaloop and its transient closing 5’-G-U-3’ base pair. This integrated approach is a critical step towards a more complete biophysical characterisation of large RNA apical loops beyond tetra and pentaloops. Our deconvolution of the ensemble into conformations defined by specific interactions and dynamics provides a basis for future ensemble-functional characterisation of this conserved non-coding RNA element and its mutants occurring in variants of concern of SARS-CoV-2.
19 3 - Research DataNMR characterisation of the antibiotic resistance-mediating 32mer RNA from the 23S ribosomal RNA2025-02-26The increasing prevalence of antibiotic resistance represents a significant public health concern, underscoring the urgent need for the development of novel therapeutic strategies. Macrolides, the second most widely used class of antibiotics, are inhibited by Erm-proteins through the methylation of adenosine 2058 of the 23S ribosomal RNA (rRNA) (~2900 nucleotides). This methylation is the molecular basis for preventing macrolides from binding and bacteria (Staphylococcus, Streptococcus and Enterococcus) developing resistance. While Erm-proteins have received considerable attention, the role of the ribosomal RNA in acquiring antibiotic resistance is frequently undervalued, even though the ribosomal RNA is the chemical target for methylation. Here, we present the comprehensive resonance assignment for 1H, 13C and 15N for the part of the 23S RNA that serves as the Erm substrate in antimicrobial resistance by solution NMR spectroscopy. Furthermore, we compare the chemical shift signature of the methylated and dimethylated RNA construct and show that changes in the RNA upon methylation are locally restricted. The chemical shift assignments provide a starting point for investigating and targeting the molecular mechanism of the resistance-conferring Erm proteins.
14 - Research DataPELDOR on multi-nitroxide model compounds2025-01-31We investigated the accuracy and limitation of using the modulation depth of pulsed electron-electron double resonance experiments to count the number of coupled spins. For this purpose synthesized multi-nitroxide molecules with two to six spins were used. We could show that the main limitation on accurately counting larger number of coupled spins at Q-band frequencies is determined by the reproducibility of adjusting and calibrating the pump pulse excitation efficiency. Contrariwise, with broadband sech/tanh or short 10 ns rectangular pump pulses modulation depth suppression effects arising from non-ideal coverage of the dipolar-split signals can be avoided for molecules with intra-molecular spin distances larger than 2nm. The transverse relaxation times for our model compounds with one to six spins did not depend on the spin number and were all the same. Nevertheless, the signal intensity of the primary Hahn echo signal in a 4-pulse PELDOR sequence decreased strongly with the number of coupled spins. This is due to the dipolar defocusing if more than one spin is excited by the first two pulses at the detection frequency, resulting in a loss of refocused echo intensity of the PELDOR experiments. This effect further reduces the accuracy of using the PELDOR modulation depth for spin counting. Altogether, our results demonstrate, that this method can potentially be applied for application up to hexameric complexes.
23 4 - Research DataExploring the modulation of the complex folding landscape of Human Telomeric DNA by low molecular weight ligands2025-02-04Telomeric DNA forms G-quadruplex (G4) structures. G4s are crucial for genomic stability and therapeutic targeting. Using time-resolved NMR and CD spectroscopies, we investigated how the ligand Phen-DC3 modulates the folding of the human telomeric repeat 23TAG DNA sequence into G4. The kinetics are modulated by the ligand and by the presence of potassium cations (K+). Ligand binding to G4 occurs via a triphasic process with fast and slow phases. Notably, for the G4 structure in the presence of K+, the slow rate is ten times slower than without K+. These findings offer key insights into the modulation of the complex folding landscape of G4s by ligands, advancing our understanding of G4-ligand interactions for potential therapeutic applications.
5 54 - Research DataEnergy Transfer Booster: How a Leaving group controls the excited state pathway within a caging BASHY-BODIPY dyad - All spectrocopic and theoretical data2024-12-19All spectroscopic and theoretical data from the manuscript are available.
45 4 - Research DataStructural response of G protein binding to the cyclodepsipeptide inhibitor FR900359 probed by NMR spectroscopy2024-07-04The cyclodepsipeptide FR900359 (FR) and its analogs are able to selectively inhibit the class of Gq proteins by blocking GDP/GTP exchange. The inhibitor binding site of Gq has been characterized by X-ray crystallography, and various binding and functional studies have determined binding kinetics and mode of inhibition. Here we investigate isotope-labeled FR bound to the membrane-anchored G protein heterotrimer by solid-state nuclear magnetic resonance (ssNMR) and in solution by liquid-state NMR. The resulting data allowed us to identify regions of the inhibitor which show especially pronounced effects upon binding and revealed a generally rigid binding mode in the cis conformation under native-like conditions. The inclusion of the membrane environment allowed us to show a deep penetration of FR into the lipid bilayer illustrating a possible access mode of FR into the cell. Dynamic nuclear polarization (DNP)-enhanced ssNMR was used to observe the structural response of specific segments of the Gα subunit to inhibitor binding. This revealed rigidification of the switch I binding site and an allosteric response in the α5 helix as well as suppression of structural changes induced by nucleotide exchange due to inhibition by FR. Our NMR studies of the FR-G protein complex conducted directly within a native membrane environment provide important insights into the inhibitors access via the lipid membrane, binding mode, and structural allosteric effects.
28 4 - Research DataMolecular Mechanisms and Evolutionary Robustness of a Color Switch in Proteorhodopsins2023-10-17The data set is associated with manuscript: Jiafei Mao, Xinsheng Jin, Man Shi, David Heidenreich, Lynda J. Brown, Richard C.D. Brown, Moreno Lelli, Xiao He, Clemens Glaubitz: Molecular Mechanisms and Evolutionary Robustness of a Color Switch in Proteorhodopsins; submitted to Science Advances (adj0384). It contains the NMR data from Figures 2, 3, 4, S2-15 as well as data for the bioinformatics analysis in Figues 1d, 7, S22-S24. An overview of all samples and spectra is provided in Table S1 of the manuscript.
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