Tryptophan synthase (TRPS) is a bifunctional enzyme consisting of α- and β-subunits that catalyzes the last two steps of L-tryptophan (L-Trp) biosynthesis. The first stage of the reaction at the ...β-subunit is called β-reaction stage I, which converts the β-ligand from an internal aldimine E(Ain) to an α-aminoacrylate E(A-A) intermediate. The activity is known to increase 3-10-fold upon the binding of 3-indole-D-glycerol-3'-phosphate (IGP) at the α-subunit. The effect of α-ligand binding on β-reaction stage I at the distal β-active site is not well understood despite the abundant structural information available for TRPS. Here, we investigate the β-reaction stage I by carrying out minimum-energy pathway searches based on a hybrid quantum mechanics/molecular mechanics (QM/MM) model. The free-energy differences along the pathway are also examined using QM/MM umbrella sampling simulations with QM calculations at the B3LYP-D3/aug-cc-pVDZ level of theory. Our simulations suggest that the sidechain orientation of βD305 near the β-ligand likely plays an essential role in the allosteric regulation: a hydrogen bond is formed between βD305 and the β-ligand in the absence of the α-ligand, prohibiting a smooth rotation of the hydroxyl group in the quinonoid intermediate, whereas the dihedral angle rotates smoothly after the hydrogen bond is switched from βD305-β-ligand to βD305-βR141. This switch could occur upon the IGP-binding at the α-subunit, as evidenced by the existing TRPS crystal structures.
Abstract
The spectroscopic features of protonated water species in dilute acid solutions have been long sought after for understanding the microscopic behavior of the proton in water with gas-phase ...water clusters H
+
(H
2
O)
n
extensively studied as bottom-up model systems. We present a new protocol for the calculation of the infrared (IR) spectra of complex systems, which combines the fragment-based Coupled Cluster method and anharmonic vibrational quasi-degenerate perturbation theory, and demonstrate its accuracy towards the complete and accurate assignment of the IR spectrum of the H
+
(H
2
O)
21
cluster. The site-specific IR spectral signatures reveal two distinct structures for the internal and surface four-coordinated water molecules, which are ice-like and liquid-like, respectively. The effect of inter-molecular interaction between water molecules is addressed, and the vibrational resonance is found between the O-H stretching fundamental and the bending overtone of the nearest neighboring water molecule. The revelation of the spectral signature of the excess proton offers deeper insight into the nature of charge accommodation in the extended hydrogen-bonding network underpinning this aqueous cluster.
An efficient anharmonic vibrational method is developed exploiting the locality of molecular vibration. Vibrational coordinates localized to a group of atoms are employed to divide the potential ...energy surface (PES) of a system into intra- and inter-group contributions. Then, the vibrational Schrödinger equation is solved based on a PES, in which the inter-group coupling is truncated at the harmonic level while accounting for the intra-group anharmonicity. The method is applied to a pentagonal hydrogen bond network (HBN) composed of internal water molecules and charged residues in a membrane protein, bacteriorhodopsin. The PES is calculated by the quantum mechanics/molecular mechanics (QM/MM) calculation at the level of B3LYP-D3/aug-cc-pVDZ. The infrared (IR) spectrum is computed using a set of coordinates localized to each water molecule and amino acid residue by second-order vibrational quasi-degenerate perturbation theory (VQDPT2). Benchmark calculations show that the proposed method yields the N–D/O–D stretching frequencies with an error of 7 cm–1 at the cost reduced by more than five times. In contrast, the harmonic approximation results in a severe error of 150 cm–1. Furthermore, the size of QM regions is carefully assessed to find that the QM regions should include not only the pentagonal HBN itself but also its HB partners. VQDPT2 calculations starting from transient structures obtained by molecular dynamics simulations have shown that the structural sampling has a significant impact on the calculated IR spectrum. The incorporation of anharmonicity, sufficiently large QM regions, and structural samplings are of essential importance to reproduce the experimental IR spectrum. The computational spectrum paves the way for decoding the IR signal of strong HBNs and helps elucidate their functional roles in biomolecules.
The infrared spectrum of H+(H2O)4 recently observed in a wide spectral range has shown a series of bands in a range of 1700–2500 cm–1, which can not be understood by the standard harmonic normal mode ...analysis. Here, we theoretically investigate the origin of these bands with a focus on (1) the possibility of coexistence of multiple isomers in the Eigen H3O+(H2O)3 and Zundel H5O2 +(H2O)2 forms and (2) the effect of anharmonic coupling that gives rise to nonzero intensities for overtones and combination bands. Anharmonic vibrational calculations are carried out for the Eigen and Zundel clusters by the second-order vibrational quasi-degenerate perturbation theory (VQDPT2) based on optimized coordinates. The anharmonic potential energy surface and the dipole moment surfaces are generated by a multiresolution approach combining one-dimensional (1D) grid potential functions derived from CCSD(T)-F12, 2D and 3D grid potential functions derived from B3LYP for important coupling terms, and a quartic force field derived from B3LYP for less important terms. The spectrum calculated for the Eigen cluster is in excellent agreement with the experiment, assigning the bands in the range of 1700–2500 cm–1 to overtones and combination bands of a H3O+ moiety in line with recent reports J. Phys. Chem. A 2015, 119, 9425 ; Science 2016, 354, 1131 . On the other hand, characteristic OH stretching bands of the Zundel cluster is found to be absent in the experimental spectrum. We therefore conclude that the experimental spectrum originates solely from the Eigen cluster. Nonetheless, the present calculation for the Eigen cluster poorly reproduces a band observed at 1765 cm–1. A possible nature of this band is discussed.
Quantum mechanics/molecular mechanics (QM/MM) calculations are applied for anharmonic vibrational analyses of biomolecules and solvated molecules. The QM/MM method is implemented into a molecular ...dynamics (MD) program, GENESIS, by interfacing with external electronic structure programs. Following the geometry optimization and the harmonic normal-mode analysis based on a partial Hessian, the anharmonic potential energy surface (PES) is generated from QM/MM energies and gradients calculated at grid points. The PES is used for vibrational self-consistent field (VSCF) and post-VSCF calculations to compute the vibrational spectrum. The method is first applied to a phosphate ion in solution. With both the ion and neighboring water molecules taken as a QM region, IR spectra of representative hydration structures are calculated by the second-order vibrational quasi-degenerate perturbation theory (VQDPT2) at the level of B3LYP/cc-pVTZ and TIP3P force field. A weight-average of IR spectra over the structures reproduces the experimental spectrum with a mean absolute deviation of 16 cm–1. Then, the method is applied to an enzyme, P450 nitric oxide reductase (P450nor), with the NO molecule bound to a ferric (FeIII) heme. Starting from snapshot structures obtained from MD simulations of P450nor in solution, QM/MM calculations have been carried out at the level of B3LYP-D3/def2-SVP(D). The spin state of FeIII(NO) is likely a closed-shell singlet state based on a ratio of N–O and Fe–NO stretching frequencies (νN–O and νFe–NO) calculated for closed- and open-shell singlet states. The calculated νN–O and νFe–NO overestimate the experimental ones by 120 and 75 cm–1, respectively. The electronic structure and solvation of FeIII(NO) affect the structure around the heme of P450nor leading to an increase in νN–O and νFe–NO.
Recent advances in atomistic molecular dynamics (MD) simulations of biomolecules allow us to explore their conformational spaces widely, observing large-scale conformational fluctuations or ...transitions between distinct structures. To reproduce or refine experimental data using MD simulations, structure ensembles, which are characterized by multiple structures and their statistical weights on the rugged free-energy landscapes, are often used. Here, we summarize weight average approaches for various experimental measurements. Weight average approaches are now applied to hybrid quantum mechanics/molecular mechanics MD simulations to predict fast vibrational motions in a protein with a high accuracy for better understanding of molecular functions from atomic structures.
•Conformational dynamics of biomolecules involves fast vibrational motions and slow transitions between distinct structures.•The conformational dynamics is theoretically described via multidimensional free-energy landscapes with many basins.•structural ensemble from molecular dynamics simulations is used to reproduce dynamical properties observed experimentally.•Weight average approaches are useful to analyze not only slow conformational changes but also fast vibrational motions.
Fluorogenic probes for bioimaging have become essential tools for life science and medicine, and the key to their development is a precise understanding of the mechanisms available for fluorescence ...off/on control, such as photoinduced electron transfer (PeT) and Förster resonance energy transfer (FRET). Here we establish a new molecular design strategy to rationally develop activatable fluorescent probes, which exhibit a fluorescence off/on change in response to target biomolecules, by controlling the twisted intramolecular charge transfer (TICT) process. This approach was developed on the basis of a thorough investigation of the fluorescence quenching mechanism of N-phenyl rhodamine dyes (commercially available as the QSY series) by means of time-dependent density functional theory (TD-DFT) calculations and photophysical evaluation of their derivatives. To illustrate and validate this TICT-based design strategy, we employed it to develop practical fluorogenic probes for HaloTag and SNAP-tag. We further show that the TICT-controlled fluorescence off/on mechanism is generalizable by synthesizing a Si–rhodamine-based fluorogenic probe for HaloTag, thus providing a palette of chemical dyes that spans the visible and near-infrared range.
A circularly polarized ultraviolet (UV) laser pulse may excite a unidirectional valence-type electronic ring current in an oriented molecule, within the pulse duration of a few femtoseconds (e.g., τ ...= 3.5 fs). The mechanism is demonstrated by quantum model simulation for |X〉 = |1 1A1g〉 → |E+〉 = |4 1Eu+〉 population transfer in the model system, Mg−porphyrin. The net ring current generated by the laser pulse (I = 84.5 μA) is at least 100 times stronger than any ring current, which could be induced by means of permanent magnetic fields, with present technology.
Reverse osmosis membranes based on aromatic polyamide (ar-PA) are widely used in desalination of seawater, yet the microscopic mechanism of water diffusion through a polyamide layer remains elusive. ...Here, we study the structure and dynamics of polymer chains and water molecules in ar-PA in comparison to nylon 6 (one of aliphatic polyamides) under various water contents (0.0–15.9 wt%). The infrared (IR) difference spectrum between dry and moist ar-PA shows little change in amide A bands, in contrast to that of nylon 6 which yields a prominent dip. Theoretical analyses using molecular dynamics simulations and quantum electronic and vibrational calculations reveal that the dip in nylon 6 is caused by breaking of hydrogen bonds (HBs) among amide groups. The incoming water molecules that break amide-amide HBs are bound to polyamide chains nearby and diffuse slowly. On the other hand, the amide-amide HBs of ar-PA are kept upon hydration. Such polymer structure facilitates growth of large water clusters with more than 100 water molecules and rapid diffusion of water molecules. The amide A band serves as a fingerprint to characterize the water permeability of polyamide materials.
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•A dip of amide A, present in nylon 6, disappears in RO (aromatic polyamides).•The absence indicates that amide-amide hydrogen bonds (HBs) are kept upon hydration.•The HB structure facilitates growth of water clusters and rapid water diffusion.•The amide A band is a fingerprint to probe the water permeability.
Structures of polyamide 6 are investigated for different hydration levels using molecular dynamics (MD) simulations and quantum vibrational calculations. The MD simulations have shown that hydration ...leads to an increase in the diffusion coefficient, accompanied by a growth of water clusters in the polymer. The IR difference spectra upon hydration are calculated using a weight-averaged method incorporating anharmonicity of the potential energy surface. The predicted IR difference spectrum for the amide A band is in quantitative agreement with the experiment Iwamoto, R. ; Murase, H. J. Polym. Sci., Part B: Polym. Phys. 2003, 41, 1722−1729 . The proposed method, combined with experimental IR difference spectra, makes it feasible to elucidate the atomistic structure of hydrated polymer materials.
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