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Uncertain chemical mechanisms leading to brown carbon (BrC) formation affect the drivers of the radiative effects of aerosols in current climate predictions. Herein, the aqueous-phase reactions of methylglyoxal (MG) and typical reduced nitrogen species (RNSs) are systematically investigated by using combined quantum chemical calculations and laboratory experiments. Imines and diimines are identified from the mixtures of methylamine (MA) and ammonia (AM) with MG, but not from dimethylamine (DA) with the MG mixture under acidic conditions, because deprotonation of DA cationic intermediates is hindered by the amino groups occupied by two methyl groups. It leads to N-heterocycle (NHC) formation in the MG + MA (MGM) and MG + AM (MGA) reaction systems but to N-containing chain oligomer formation in the MG + DA (MGD) reaction system. Distinct product formation is attributed to electrostatic attraction and steric hindrance, which are regulated by the methyl groups of RNSs. The light absorption and adverse effects of NHCs are also strongly related to the methyl groups of RNSs. Our finding reveals that BrC formation is mainly contributed from MG reaction with RNSs with less methyl groups, which have more abundant and broad sources in the urban environments.

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Developing functionally complex carbon materials from small aromatic molecules requires an understanding of how the chemistry and structure of its constituent molecules evolve and crosslink, to achieve a tailorable set of functional properties. Here, molecular dynamics (MD) simulations are used to isolate the effect of methyl groups on condensation reactions during the oxidative process and evaluate the impact on elastic modulus by considering three monodisperse pyrene-based systems with increasing methyl group fraction. A parameter to quantify the reaction progression is designed by computing the number of new covalent bonds formed. Utilizing the previously developed MD framework, it is found that increasing methylation leads to an almost doubling of bond formation, a larger fraction of the new bonds oriented in the direction of tensile stress, and a higher basal plane alignment of the precursor molecules along the direction of tensile stress, resulting in enhanced tensile modulus. Additionally, via experiments, it is demonstrated that precursors with a higher fraction of methyl groups result in a higher alignment of molecules. Moreover, increased methylation results in the lower spread of single molecule alignment which may lead to smaller variations in tensile modulus and more consistent properties in carbon materials derived from methyl-rich precursors.

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Protein methyl groups can participate in multiple motional modes on different time scales. Sub-nanosecond to nano-second time scale motions of methyl axes are particularly challenging to detect for small proteins in solutions. In this work we employ NMR relaxation interference between the methyl H-H/H-C dipole-dipole interactions [Sun&Tugarinov, J. Magn. Reason. 2012] to characterize methyl axes motions as a function of temperature in a small model protein villin headpiece subdomain (HP36), in which all non-exchangeable protons are deuterated with the exception of methyl groups of leucine and valine residues. The data points to the existence of slow motional modes of methyl axes on sub-nanosecond to nanosecond time scales. Further, at high temperatures for which the overall tumbling of the protein is on the order of 2 ns, we observe a coupling between the slow internal motion and the overall molecular tumbling, based on the anomalous order parameters and their temperature-dependent trends. The addition of 28%(w/w) glycerol-d8 increases the viscosity of the solvent and separates the timescales of internal and overall tumbling, thus permitting for another view of the necessity of the coupling assumption for these sites at high temperatures.

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HSP90 is a major molecular chaperone that helps both folding and stabilization of various client proteins often implicated in growth control and cell survival such as kinases and transcription factors. However, among HSP90 clients are also found numerous oncoproteins and, through its assistance to them, HSP90 has consequently been reported as a promising anticancer target. Several ligand chemotypes, including resorcinol type ligands, were found to inhibit HSP90, most of them in an ATP competitive manner. Binding of some of these ligands modify significantly the NMR spectrum of the HSP90 ATP binding domain compared to the apo protein spectrum, hampering assignment transfer from the previously assigned human HSP90 apo state. Here we report the assignment of the 1HN, 15N, 13C', 13Cα, 13Cß, 1Hmethyl, and 13Cmethyl chemical shifts of the 29 kDa HSP90 N-terminal domain bound to a long residence time resorcinol type inhibitor: 5-[4-(2-Fluoro-phenyl)-5-oxo-4,5-dihydro-1H-[1,2,4]triazol-3-yl]-N-furan-2-ylmethyl-2,4-dihydroxy-N-methyl-benzamide. 92% of the backbone resonances and 100% of the [1H, 13C]-resonances of Aß, Mε, Tγ, Lδ2, Vγ2 and Iδ1 methyl groups were successfully assigned, including for the first time the assignment of the segment covering the nucleotide/drug binding site. Secondary structure predictions based on the NMR assignment reveal a structural rearrangement of HSP90 N-terminal domain upon ligand binding. The long residence time ligand induces the formation of a continuous helix covering the ligand binding site of HSP90 N-terminal domain accounting for the large differences observed in the NMR spectra between the apo and bound proteins.

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In nuclear magnetic resonance spectroscopy of proteins, methyl protons play a particular role as extremely sensitive reporters on dynamics, allosteric effects, and protein-protein interactions, accessible even in high-molecular-weight systems approaching 1 MDa. The notorious issue of their chemical shift assignment is addressed here by a joint use of solid-state 1H-detected methods at very fast (nearly 100 kHz) magic-angle spinning, partial deuteration, and high-magnetic fields. The suitability of a series of RF schemes is evaluated for the efficient coherence transfer across entire 13C side chains of methyl-containing residues, which is key for establishing connection between methyl and backbone 1H resonances. The performance of ten methods for recoupling of either isotropic 13C-13C scalar or anisotropic dipolar interactions (five variants of TOBSY, FLOPSY, DIPSI, WALTZ, RFDR, and DREAM) is evaluated experimentally at two state-of-the-art magic-angle spinning (55 and 94.5 kHz) and static magnetic field conditions (18.8 and 23.5 T). Model isotopically labeled compounds (alanine and Met-Leu-Phe tripeptide) and ILV-methyl and amide-selectively protonated, and otherwise deuterated chicken α-spectrin SH3 protein are used as convenient reference systems. Spin dynamics simulations in SIMPSON are performed to determine optimal parameters of these RF schemes, up to recently experimentally attained spinning frequencies (200 kHz) and B 0 field strengths (28.2 T). The concept of linearization of 13C side chain by appropriate isotope labeling is revisited and showed to significantly increase sensitivity of methyl-to-backbone correlations. A resolution enhancement provided by 4D spectroscopy with non-uniform (sparse) sampling is demonstrated to remove ambiguities in simultaneous resonance assignment of methyl proton and carbon chemical shifts.

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Methyl moieties are highly valuable probes for quantitative NMR studies of large proteins. Hence, their assignment is of the utmost interest to obtain information on both interactions and dynamics of proteins in solution. Here, we present the synthesis of a new precursor that allows connection of leucine and valine pro-S methyl moieties to backbone atoms by linear 13C-chains. This optimized 2H/13C-labelled acetolactate precursor can be combined with existing 13C/2H-alanine and isoleucine precursors in order to directly transfer backbone assignment to the corresponding methyl groups. Using this simple approach leucine and valine pro-S methyl groups can be assigned using a single sample without requiring correction of 1H/2H isotopic shifts on 13C resonances. The approach was demonstrated on the N-terminal domain of human HSP90, for which complete assignment of Ala-ß, Ile-δ1, Leu-δ2, Met-ε, Thr-γ and Val-γ2 methyl groups was obtained.

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For effective removal of cadmium (II) (Cd(II)) from polluted water, a magnetic adsorbent of Fe3O4@SiO2 core-shell microspheres modified with methyl-protected thiol groups (Fe3O4@SiO2-SH-Protected) was synthesized and characterized by scanning electron, transmission electron, energy dispersive X-ray spectroscopy, Fourier transform infrared spectroscopies, as well as X-ray diffraction, Raman spectroscopy, and magnetic measurements. Characterization results showed that thiol groups on the surface of Fe3O4@SiO2 material were protected to avoid disulfide formation. Batch adsorption experiments were conducted by varying the contact time, initial pH, solid-liquid ratio, temperature, Cd(II) concentrations, and interfering cations. Fe3O4@SiO2-SH-Protected material exhibited much higher adsorption capacity than Unprotected forms and other adsorbents due to methyl group protection. The maximum adsorption capacity calculated from the Langmuir fitting was 27.5 mg·g-1 (pH 7, 25 °C), and the adsorption kinetics followed a pseudo-second-order model, and adsorption mainly dominated by film diffusion processes. Thermodynamic parameters indicated that the adsorption process was a spontaneous, endothermic, and positive entropic process. Cd(II)-loaded on the adsorbent was easily desorbed with 0.1 M HCl and the adsorbent stable in 0.1 M HCl for long times, showing good reusability and stability.

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The properties of graphdiyne (GDY), such as energy gap, morphology, and affinity to alkali metals, can be adjusted by including electron-withdrawing/donating groups. The push-pull electron ability and size differences of groups play a key role on the partial property adjusting of GDY derivatives MeGDY, HGDY, and CNGDY. Cyano groups (electron-withdrawing) and methyl groups (electron-donating) decrease the band gap and increase the conductivity of the GDY network. The cyano and methyl groups affects the aggregation of GDY, providing a higher number of micropores and specific surface area. These groups also endow the original GDY additional advantages: the stronger electronegativity of cyano groups increase the affinity of GDY frameworks to lithium atoms, and the larger atomic volume of methyl groups increases the interlayer distance and provides more storage space and diffusion tunnels.

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A major goal in structural biology is to unravel how molecular machines function in detail. To that end, solution-state NMR spectroscopy is ideally suited as it is able to study biological assemblies in a near natural environment. Based on methyl TROSY methods, it is now possible to record high-quality data on complexes that are far over 100 kDa in molecular weight. In this review, we discuss the theoretical background of methyl TROSY spectroscopy, the information that can be extracted from methyl TROSY spectra and approaches that can be used to assign methyl resonances in large complexes. In addition, we touch upon insights that have been obtained for a number of challenging biological systems, including the 20S proteasome, the RNA exosome, molecular chaperones and G-protein-coupled receptors. We anticipate that methyl TROSY methods will be increasingly important in modern structural biology approaches, where information regarding static structures is complemented with insights into conformational changes and dynamic intermolecular interactions.

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Specific isotopic labeling of methyl groups in a perdeuterated protein background enables the detection of long range NOEs in proteins or high molecular weight complexes. We introduce here an approach, combining an optimized isotopic labeling scheme with a specifically tailored NMR pulse sequence, to distinguish between intramolecular and intermolecular NOE connectivities. In hetero-oligomeric complexes, this strategy enables sign encoding of intra-subunit and inter-subunit NOEs. For homo-oligomeric assemblies, our strategy allows the specific detection of intra-chain NOEs in high resolution 3D NOESY spectra. The general principles, possibilities and limitations of this approach are presented. Applications of this approach for the detection of intermolecular NOEs in a hetero-hexamer, and the assignment of methyl 1H and 13C resonances in a homo-tetrameric protein complex are shown.

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Optimized NMR experiments are developed for isolating magnetization belonging to the I=1/2 manifolds of 13 CH3 methyl groups in proteins, enabling the manipulation of the magnetization of a 13 CH3 moiety as if it were an AX (1 H-13 C) spin-system. These experiments result in the same 'simplification' of a 13 CH3 spin-system that would be obtained from the production of {13 CHD2 }-methyl-labeled protein samples. The sensitivity of I=1/2 manifold-selection experiments is a factor of approximately 2 less than that of the corresponding experiments acquired on {13 CHD2 }-labeled methyl groups. The methodology described here is primarily intended for small-to-medium sized proteins, where the losses in sensitivity associated with the isolation of I=1/2 manifold transitions can be tolerated. Several NMR applications that benefit from simplification of the 13 CH3 (AX3 ) spin-systems are described, with an emphasis on the measurements of methyl 1 H-13 C residual dipolar couplings in a {13 CH3 }-methyl-labeled deletion mutant of the human chaperone DNAJB6b, where modulation of NMR signal intensities due to evolution of methyl 1 H-13 C scalar and dipolar couplings follows a simple cosine function characteristic of an AX (1 H-13 C) spin-system, significantly simplifying data analysis.

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Carbohydrates, also known as glycans in biological systems, are omnipresent in nature where they as glycoconjugates occur as oligo- and polysaccharides linked to lipids and proteins. Their three-dimensional structure is defined by two or three torsion angles at each glycosidic linkage. In addition, transglycosidic hydrogen bonding between sugar residues may be important. Herein we investigate the presence of these inter-residue interactions by NMR spectroscopy in D2 O/[D6 ]DMSO (70:30) or D2 O and by molecular dynamics (MD) simulations with explicit water as solvent for disaccharides with structural elements α-d-Manp-(1→2)-d-Manp, ß-d-GlcpNAc-(1→2)-d-Manp, and α-d-Glcp-(1→4)-ß-d-Glcp, all of which have been suggested to exhibit inter-residue hydrogen bonding. For the disaccharide ß-d-GlcpNAc-(1→2)-ß-d-Manp-OMe, the large extent of O5'⋅⋅⋅HO3 hydrogen bonding as seen from the MD simulation is implicitly supported by the 1 H NMR chemical shift and 3 JHO3,H3 value of the hydroxy proton. In the case of α-d-Glcp-(1→4)-ß-d-Glcp-OMe, the existence of a transglycosidic hydrogen bond O2'⋅⋅⋅HO3 was proven by the presence of a cross-peak in 1 H,13 C HSQC-TOCSY experiments as a result of direct TOCSY transfer between HO3 of the reducing end residue and H2' (detected at C2') of the terminal residue. The occurrence of inter-residue hydrogen bonding, albeit transient, is judged important for the stabilization of three-dimensional structures, which may be essential in maintaining a conformational state for carbohydrate-protein interactions of glycans to take place in biologically important environments.

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Residual dipolar couplings (RDCs) provide both structural and dynamical information useful in the characterization of biological macromolecules. While most data come from the interaction of simple pairs of directly bonded spin-1/2 nuclei (1H-15N, 1H-13C, 1H-1H), it is possible to acquire data from interactions among the multiple spins of 13C-labeled methyl groups (1H3-13C). This is especially important because of the advantages that observation of 13C-labeled methyl groups offers in working with very large molecules. Here we consider some of the options for measurement of methyl RDCs in large and often fully protonated proteins and arrive at a pulse sequence that exploits both J-modulation and direct detection of 13C. Its utility is illustrated by application to a fully protonated two domain fragment from the mammalian glycoprotein, Robo1, 13C-methyl-labeled in all valines.

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With the development of sophisticated pulsed field gradient- and phase cycling-approaches for suppressing certain coherence transfer pathways and selecting for others it is sometimes easy to forget that the process is not flawless. In some cases artifacts can emerge because unwanted transfers are immune to the phase cycle or the application of gradients. We consider here a simple 1H,13C HMQC pulse scheme and show that imperfections in the single 1H 180° refocusing pulse can give rise to small artifacts in methyl spectra that cannot be eliminated through extensive phase cycling or the use of gradients, but that are easily removed when the pulse is of the composite variety.

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A new and practical α-monomethylation strategy using an amine-borane/N,N-dimethylformamide (R3 N-BH3 /DMF) system as the methyl source was developed. This protocol has been found to be effective in the α-monomethylation of arylacetonitriles and arylacetamides. Mechanistic studies revealed that the formyl group of DMF delivered the carbon and one hydrogen atoms of the methyl group, and R3 N-BH3 donated the remaining two hydrogen atoms. Such a unique reaction pathway enabled controllable assemblies of CDH2 -, CD2 H-, and CD3 - units using Me2 NH-BH3 /d7 -DMF, Me3 N-BD3 /DMF and Me3 N-BD3 /d7 -DMF systems, respectively. Further application of this method to the facile synthesis of anti-inflammatory flurbiprofen and its varied deuterium-labeled derivatives was demonstrated.

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Liquid-state NMR spectroscopy is a powerful technique to elucidate binding properties of ligands on proteins. Ligands binding in hydrophobic pockets are often in close proximity to methyl groups and binding can lead to subtle displacements of methyl containing side chains to accommodate the ligand. To establish whether pseudocontact shifts can be used to characterize ligand binding and the effects on methyl groups, the N-terminal domain of HSP90 was tagged with caged lanthanoid NMR probe 5 at three positions and titrated with a ligand. Binding was monitored using the resonances of leucine and valine methyl groups. The pseudocontact shifts (PCS) caused by ytterbium result in enhanced dispersion of the methyl spectrum, allowing more resonances to be observed. The effects of tag attachment on the spectrum and ligand binding are small. Significant changes in PCS were observed upon ligand binding, indicating displacements of several methyl groups. By determining the cross-section of PCS iso-surfaces generated by two or three paramagnetic centers, the new position of a methyl group can be estimated, showing displacements in the range of 1-3 Å for methyl groups in the binding site. The information about such subtle but significant changes may be used to improve docking studies and can find application in fragment-based drug discovery.

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In cryogenic dissolution NMR experiments, a substance of interest is allowed to rest in a strong magnetic field at cryogenic temperature, before dissolving the substance in a warm solvent, transferring it to a high-resolution NMR spectrometer, and observing the solution-state NMR spectrum. In some cases, negative enhancements of the 13C NMR signals are observed, which have been attributed to quantum-rotor-induced polarization. We show that in the case of acetone (propan-2-one) the negative signal enhancements of the methyl 13C sites may be understood by invoking conventional cross-relaxation within the methyl groups. The 1H nuclei acquire a relative large net polarization through thermal equilibration in a magnetic field at low temperature, facilitated by the methyl rotation which acts as a relaxation sink; after dissolution, the 1H magnetization slowly returns to thermal equilibrium at high temperature, in part by cross-relaxation processes, which induce a transient negative polarization of nearby 13C nuclei. We provide evidence for this mechanism experimentally and theoretically by saturating the 1H magnetization using a radiofrequency field pulse sequence before dissolution and comparing the 13C magnetization evolution after dissolution with the results obtained from a conventional 1H-13C cross relaxation model of the CH3 moieties in acetone.

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We propose a proof-of-principle experiment to encode one logical qubit in noise protected subspace of three identical spins in a methyl group. The symmetry analysis of the wavefunction shows that this fermionic system exhibits a symmetry correlation between the spatial degree of freedom and the spin degree of freedom. We show that one can use this correlation to populate the noiseless subsystem by relying on the interaction between the electric dipole moment of the methyl group with a circularly polarized microwave field. Logical gates are implemented by controlling both the intensity and phase of the applied field.

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A new version of the program PARAssign has been evaluated for assignment of NMR resonances of the 76 methyl groups in leucines, isoleucines and valines in a 25 kDa protein, using only the structure of the protein and pseudocontact shifts (PCS) generated with a lanthanoid tag at up to three attachment sites. The number of reliable assignments depends strongly on two factors. The principle axes of the magnetic susceptibility tensors of the paramagnetic centers should not be parallel so as to avoid correlated PCS. Second, the fraction of resonances in the spectrum of a paramagnetic sample that can be paired with the diamagnetic counterparts is critical for the assignment. With the data from two tag positions a reliable assignment could be obtained for 60% of the methyl groups and for many of the remaining resonances the number of possible assignments is limited to two or three. With a single tag, reliable assignments can be obtained for methyl groups with large PCS near the tag. It is concluded that assignment of methyl group resonances by paramagnetic tagging can be particularly useful in combination with some additional data, such as from mutagenesis or NOE-based experiments. Approaches to yield the best assignment results with PCS generating tags are discussed.

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Many O-methyl and N-methyl groups in natural products are depleted in 13C relative to the rest of the molecule. These methyl groups are derived from the C-1 tetrahydrofolate pool via l-methionine, the principle donor of methyl units. Depletion could occur at a number of steps in the pathway. We have tested the hypothesis that methionine biosynthesis is implicated in this depletion by using a combined experimental and theoretical approach. By using isotope ratio monitoring 13C NMR spectrometry to measure the position-specific distribution of 13C within l-methionine of natural origin, it is shown that the S-methyl group is depleted in 13C by ∼20‰ relative to the other positions in the molecule. In parallel, we have conducted a basic theoretical analysis of the reaction pathway of methionine synthase to assess whether the enzyme cobalamin-independent l-methionine synthase (EC 2.1.1.14)-that catalyzes the synthesis of l-methionine from 5-methyl-tetrahydrofolate and homocysteine-plays a role in causing this depletion. Calculation predicts a strong normal 13C kinetic isotope effect (1.087) associated with this enzyme. Hence, depletion in 13C in the S-methyl of l-methionine during biosynthesis can be identified as an important factor contributing to the general depletion seen in many O-methyl and N-methyl groups of natural products.

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