Identification of allosteric hotspots regulating the ribosomal RNA binding by antibiotic resistance-conferring Erm methyltransferases

Antibiotic resistance via epigenetic methylation of ribosomal RNA is one of the most prevalent strategies adopted by multidrug resistant pathogens. The erythromycin-resistance methyltransferase (Erm) methylates rRNA at the conserved A2058 position and imparts resistance to macrolides such as erythromycin. However, the precise mechanism adopted by Erm methyltransferases for locating the target base within a complicated rRNA scaffold remains unclear. Here, we show that a conserved RNA architecture, including specific bulge sites, present more than 15 Å from the reaction center, is key to methylation at the pathogenic site. Using a set of RNA sequences site-specifically labeled by fluorescent nucleotide surrogates, we show that base flipping is a prerequisite for effective methylation and that distal bases assist in the recognition and flipping at the reaction center. The Erm–RNA complex model revealed that intrinsically flipped-out bases in the RNA serve as a putative anchor point for the Erm. Molecular dynamic simulation studies demonstrated the RNA undergoes a substantial change in conformation to facilitate an effective protein–rRNA handshake. This study highlights the importance of unique architectural features exploited by RNA to impart fidelity to RNA methyltransferases via enabling allosteric crosstalk. Moreover, the distal trigger sites identified here serve as attractive hotspots for the development of combination drug therapy aimed at reversing resistance

Role of Alkyl Chain Linker in the Conformational Preferences of N-phenylalkylsquaramates and bis-N,N\u27-diphenylalkylsquaramides

The conformational space of small molecules can be exploited to deploy the hierarchical nature of the intermolecular interactions such as hydrogen bonding and π-stacking in the crystal structures. Modulation of the conformational space was achieved by varying the alkyl chain length in N-phenylalkylsquaramates and bis-N,N\u27-diphenylalkylsquaramides, which were synthesized by condensation of dimethyl squarate with the corresponding phenyl alkylamines. In the crystal structures of these compounds, even though the catemeric N–H···O hydrogen bonding is the primary interaction and the hierarchy in the secondary interactions consisting of π-stacking and H···H bonding leads to a large variation in the molecular structures in the solid state

Vibrational Stark Fields in Carboxylic Acid Dimers

Carboxylic acids form exceptionally stable dimers and have been used to model proton and double proton transfer processes. The stabilization energies of the carboxylic acid dimers are very weakly dependent on the nature of the substitution. However, the electric field experienced by the OH group of a particular carboxylic acid is dependent more on the nature of the substitution on the dimer partner. In general, the electric field was higher when the partner was substituted with electron-donating group and lower with electron-withdrawing substituent on the partner. The Stark tuning rate (∆μ) of the O–H stretching vibrations calculated at the MP2/aug-cc-pVDZ level was found to be weakly dependent on the nature of substitution on the carboxylic acid. The average Stark tuning rate of O–H stretching vibrations of a particular carboxylic acid when paired with other acids was 5.7 cm–1 (MV cm–1)–1, while the corresponding average Stark tuning rate of the partner acids due to a particular carboxylic acid was 21.9 cm–1 (MV cm–1)–1. The difference in the Stark tuning rate is attributed to the primary and secondary effects of substitution on the carboxylic acid. The average Stark tuning rate for the anharmonic O–D frequency shifts is about 40-50% higher than the corresponding harmonic O–D frequency shifts calculated at B3LYP/aug-cc-pVDZ level, much greater than the typical scaling factors used, indicating the strong anharmonicity of O–H/O–D oscillators in carboxylic acid dimers. Finally, the linear correlation observed between pKa and the electric field was used to estimate the pKa of fluoroformic acid to be around 0.9

CYCLOHEXANE AS AN IONOPHORE

Author Institution: Department of Chemistry, University of Illinois at Urbana ChampaignSeveral molecules based upon 1,3,5-triaxially substituted cyclohexane templates have shown promising results as ionophores. More importantly, in the case of $Li^{+}$, cyclohexane is a more selective ionophore than the traditional 12-Crown-4 macrocyclic ether. In order to understand the effect of the alkali metal ion binding to cyclohexane, the infrared spectra of $Li^{+}$, $Na^{+}$, and $K^{+}$ ions complexed with cyclohexane were recorded in C-H stretching region. The effect of the ion binding on the C-H stretching vibrations will be discussed

VIBRONIC SPECTROSCOPY AND IVR DYNAMICS IN THE S1S_{1} STATE OF JET COOLED pp-ALKOXYANILINES; (NH2−C6H4−OCnH2n+1m=4,5,6)(NH_{2}-C_{6}H_{4}-OC_{n}H_{2n+1} m= 4,5,6)

$^{a}$S. Wategoankar and S. Doraiswamy, J. Chem. Phys. 106, in press (1997).Author Institution: Chemical Physics Group, Tata Institute of Fundamental ResearchThe ($S_{1} \leftarrow S_{0}$) excitation and the single vibronic level fluorescence (SVLF) spectra of p-butoxyaniline, p-pentyloxyaniline, and p-hexyloxyaniline were recorded in a supersonic jet expansion apparatus. The band origin transitions for each of the compounds indicated the presence of multiple geometric isomers. The SVLF spectra will be discussed vis-a-vis the existence of the isomers of each of these, and the IVR dynamics in the p-alkoxyaniline series. The IVR dynamics will be compared with that $observed^{a}$ in the first three members of the series viz. p-aminophenol, p-methoxyaniline, and p-ethoxyaniline

Unraveling the Significance of Mg²⁺ Dependency and Nucleotide Binding Specificity of H‑RAS

RAS is a small GTPase and acts as a binary molecular switch; the transition from its active to inactive state plays a crucial role in various cell signaling processes. Molecular dynamics simulations at the atomistic level suggest that the absence of cofactor Mg2+ ion generally leads to pronounced structural changes in the Switch-I than Switch-II regions and assists GTP binding. The presence of the Mg2+ ion also restricts the rotation of ϒ phosphate and enhances the hydrolysis rate of GTP. Further, the simulations reveal that the stability of the protein is almost uncompromised when Mg2+ is replaced with Zn2+ and not the Ca2+ ion. The specificity of H-RAS to GTP was evaluated by substituting with ATP and CTP, which indicates that the binding pocket tolerates purine bases over pyrimidine bases. However, the D119 residue specifically interacts with the guanine base and serves as one of the primary interactions that leads to the selectivity of GTP over ATP. The ring displacement of 32Y serves as gate dynamics in H-RAS which are important for its interaction with GAP for the nucleotide exchange and is restricted in the presence of ATP. Finally, the point mutations 61, 16, and 32 influence the structural changes, specifically in the Switch-II region, which are expected to impact the GTP hydrolysis and thus are termed oncogenic mutations

FRANCK-CONDON SPECTRAL CALCULATION ON TRANS-HYDROQUIONE USING CORRELATION FUNCTION APPROACH

Author Institution: Department of Chemical Sciences, Tata Institute of Fundamental Research; School of Chemistry, University of HyderabadIVR in polyatomic molecules has been found to be extremely fast even at energies as low as a few hundred wavenumbers. In order to understand the IVR dynamics better one can porceed to calculate the fluorescence spectra and compare with the experimental data. However, for polyatomic molecules with large number of vibrational modes even calculating a simple Franck-Condon spectrum is not a simple task. As a first step towards this we have calculated the excitation and dispersed fluorescence spectra of hydroquinone, ab initio, using a correlation function approach using a quadratic Hamiltonian. Time dependent coupled cluster method was used to propogate the wavefunction in time. The comparison between the calculated and experimental spectra will be presented and the validity of the method will be discussed $vis-\'{a}-vis$ the observed IVR behaviour