Ab initio calculation of electronic coupling in the photosynthetic reaction center

We have carried out an ab initio electronic structure calculations of electron transfer couplings between chromophores in the bacterial photosynthetic reaction center. The couplings agree remarkably well with parameters obtained from recent quantum dynamical modeling of experimental data assuming an explicit intermediate mechanism. We also have computed couplings on the M-side of the reaction center and have found that the interaction of the primary donor to the M-side intermediate bacteriochlorophyll is quite small because of destructive interference of the two localized coupling matrix elements. This may explain the slow rate of electron transfer down the M-side of the reaction center.

Molecular dynamics of a grid-mounted molecular dipolar rotor in a rotating electric field

Classical molecular dynamics is applied to the rotation of a dipolar molecular rotor mounted on a square grid and driven by rotating electric field E(ν) at T ≃ 150 K. The rotor is a complex of Re with two substituted o-phenanthrolines, one positively and one negatively charged, attached to an axial position of Rh\documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{document} \begin{equation*}{\mathrm{_{2}^{4+}}}\end{equation*}\end{document} in a [2]staffanedicarboxylate grid through 2-(3-cyanobicyclo[1.1.1]pent-1-yl)malonic dialdehyde. Four regimes are characterized by a, the average lag per turn: (i) synchronous (a < 1/e) at E(ν) = |E(ν)| > Ec(ν) [Ec(ν) is the critical field strength], (ii) asynchronous (1/e < a < 1) at Ec(ν) > E(ν) > Ebo(ν) > kT/μ, [Ebo(ν) is the break-off field strength], (iii) random driven (a ≃ 1) at Ebo(ν) > E(ν) > kT/μ, and (iv) random thermal (a ≃ 1) at kT/μ > E(ν). A fifth regime, (v) strongly hindered, W > kT, Eμ, (W is the rotational barrier), has not been examined. We find Ebo(ν)/kVcm−1 ≃ (kT/μ)/kVcm−1 + 0.13(ν/GHz)1.9 and Ec(ν)/kVcm−1 ≃ (2.3kT/μ)/kVcm−1 + 0.87(ν/GHz)1.6. For ν > 40 GHz, the rotor behaves as a macroscopic body with a friction constant proportional to frequency, η/eVps ≃ 1.14 ν/THz, and for ν < 20 GHz, it exhibits a uniquely molecular behavior.

Comparison of the early stages of forced unfolding for fibronectin type III modules

The structural changes accompanying stretch-induced early unfolding events were investigated for the four type III fibronectin (FN-III) modules, FN-III7, FN-III8, FN-III9, and FN-III10 by using steered molecular dynamics. Simulations revealed that two main energy barriers, I and II, have to be overcome to initiate unraveling of FN-III's tertiary structure. In crossing the first barrier, the two opposing β-sheets of FN-III are rotated against each other such that the β-strands of both β-sheets align parallel to the force vector (aligned state). All further events in the unfolding pathway proceed from this intermediate state. A second energy barrier has to be overcome to break the first major cluster of hydrogen bonds between adjacent β-strands. Simulations revealed that the height of barrier I varied significantly among the four modules studied, being largest for FN-III7 and lowest for FN-III10, whereas the height of barrier II showed little variation. Key residues affecting the mechanical stability of FN-III modules were identified. These results suggest that FN-III modules can be prestretched into an intermediate state with only minor changes to their tertiary structures. FN-III10, for example, extends 12 Å from the native “twisted” to the intermediate aligned state, and an additional 10 Å from the aligned state to further unfolding where the first β-strand is peeled away. The implications of the existence of intermediate states regarding the elasticity of fibrillar fibers and the stretch-induced exposure of cryptic sites are discussed.

Quantification of the hydrophobic interaction by simulations of the aggregation of small hydrophobic solutes in water

The hydrophobic interaction, the tendency for nonpolar molecules to aggregate in solution, is a major driving force in biology. In a direct approach to the physical basis of the hydrophobic effect, nanosecond molecular dynamics simulations were performed on increasing numbers of hydrocarbon solute molecules in water-filled boxes of different sizes. The intermittent formation of solute clusters gives a free energy that is proportional to the loss in exposed molecular surface area with a constant of proportionality of 45 ± 6 cal/mol⋅Å2. The molecular surface area is the envelope of the solute cluster that is impenetrable by solvent and is somewhat smaller than the more traditional solvent-accessible surface area, which is the area transcribed by the radius of a solvent molecule rolled over the surface of the cluster. When we apply a factor relating molecular surface area to solvent-accessible surface area, we obtain 24 cal/mol⋅Å2. Ours is the first direct calculation, to our knowledge, of the hydrophobic interaction from molecular dynamics simulations; the excellent qualitative and quantitative agreement with experiment proves that simple van der Waals interactions and atomic point-charge electrostatics account for the most important driving force in biology.

Thermodynamic parameters for the helix-coil transition of oligopeptides: molecular dynamics simulation with the peptide growth method.

The helix-coil transition equilibrium of polypeptides in aqueous solution was studied by molecular dynamics simulation. The peptide growth simulation method was introduced to generate dynamic models of polypeptide chains in a statistical (random) coil or an alpha-helical conformation. The key element of this method is to build up a polypeptide chain during the course of a molecular transformation simulation, successively adding whole amino acid residues to the chain in a predefined conformation state (e.g., alpha-helical or statistical coil). Thus, oligopeptides of the same length and composition, but having different conformations, can be incrementally grown from a common precursor, and their relative conformational free energies can be calculated as the difference between the free energies for growing the individual peptides. This affords a straightforward calculation of the Zimm-Bragg sigma and s parameters for helix initiation and helix growth. The calculated sigma and s parameters for the polyalanine alpha-helix are in reasonable agreement with the experimental measurements. The peptide growth simulation method is an effective way to study quantitatively the thermodynamics of local protein folding.

Site-specific conformational determination in thermal unfolding studies of helical peptides using vibrational circular dichroism with isotopic substitution

Understanding the detailed mechanism of protein folding requires dynamic, site-specific stereochemical information. The short time response of vibrational spectroscopies allows evaluation of the distribution of populations in rapid equilibrium as the peptide unfolds. Spectral shifts associated with isotopic labels along with local stereochemical sensitivity of vibrational circular dichroism (VCD) allow determination of the segment sequence of unfolding. For a series of alanine-rich peptides that form α-helices in aqueous solution, we used isotopic labeling and VCD to demonstrate that the α-helix noncooperatively unwinds from the ends with increasing temperature. For these blocked peptides, the C-terminal is frayed at 5°C. Ab initio level theoretical simulations of the IR and VCD band shapes are used to analyze the spectra and to confirm the conformation of the labeled components. The VCD signals associated with the labeled residues are amplified by coupling to the nonlabeled parts of the molecule. Thus small labeled segments are detectable and stereochemically defined in moderately large peptides in this report of site-specific peptide VCD conformational analysis.

Mapping the early steps in the pH-induced conformational conversion of the prion protein

Under certain conditions, the prion protein (PrP) undergoes a conformational change from the normal cellular isoform, PrPC, to PrPSc, an infectious isoform capable of causing neurodegenerative diseases in many mammals. Conversion can be triggered by low pH, and in vivo this appears to take place in an endocytic pathway and/or caveolae-like domains. It has thus far been impossible to characterize the conformational change at high resolution by experimental methods. Therefore, to investigate the effect of acidic pH on PrP conformation, we have performed 10-ns molecular dynamics simulations of PrPC in water at neutral and low pH. The core of the protein is well maintained at neutral pH. At low pH, however, the protein is more dynamic, and the sheet-like structure increases both by lengthening of the native β-sheet and by addition of a portion of the N terminus to widen the sheet by another two strands. The side chain of Met-129, a polymorphic codon in humans associated with variant Creutzfeldt–Jakob disease, pulls the N terminus into the sheet. Neutralization of Asp-178 at low pH removes interactions that inhibit conversion, which is consistent with the Asp-178–Asn mutation causing human prion diseases.

Protein folding from a highly disordered denatured state: The folding pathway of chymotrypsin inhibitor 2 at atomic resolution

Previous experimental and theoretical studies have produced high-resolution descriptions of the native and folding transition states of chymotrypsin inhibitor 2 (CI2). In similar fashion, here we use a combination of NMR experiments and molecular dynamics simulations to examine the conformations populated by CI2 in the denatured state. The denatured state is highly unfolded, but there is some residual native helical structure along with hydrophobic clustering in the center of the chain. The lack of persistent nonnative structure in the denatured state reduces barriers that must be overcome, leading to fast folding through a nucleation–condensation mechanism. With the characterization of the denatured state, we have now completed our description of the folding/unfolding pathway of CI2 at atomic resolution.

The arginine finger of RasGAP helps Gln-61 align the nucleophilic water in GAP-stimulated hydrolysis of GTP

The Ras family of GTPases is a collection of molecular switches that link receptors on the plasma membrane to signaling pathways that regulate cell proliferation and differentiation. The accessory GTPase-activating proteins (GAPs) negatively regulate the cell signaling by increasing the slow intrinsic GTP to GDP hydrolysis rate of Ras. Mutants of Ras are found in 25–30% of human tumors. The most dramatic property of these mutants is their insensitivity to the negative regulatory action of GAPs. All known oncogenic mutants of Ras map to a small subset of amino acids. Gln-61 is particularly important because virtually all mutations of this residue eliminate sensitivity to GAPs. Despite its obvious importance for carcinogenesis, the role of Gln-61 in the GAP-stimulated GTPase activity of Ras has remained a mystery. Our molecular dynamics simulations of the p21ras–p120GAP–GTP complex suggest that the local structure around the catalytic region can be different from that revealed by the x-ray crystal structure. We find that the carbonyl oxygen on the backbone of the arginine finger supplied in trans by p120GAP (Arg-789) interacts with a water molecule in the active site that is forming a bridge between the NH2 group of the Gln-61 and the γ-phosphate of GTP. Thus, Arg-789 may play a dual role in generating the nucleophile as well as stabilizing the transition state for P—O bond cleavage.

Direct evidence for modified solvent structure within the hydration shell of a hydrophobic amino acid.

Neutron scattering experiments are used to determine scattering profiles for aqueous solutions of hydrophobic and hydrophilic amino acid analogs. Solutions of hydrophobic solutes show a shift in the main diffraction peak to smaller angle as compared with pure water, whereas solutions of hydrophilic solutes do not. The same difference for solutions of hydrophobic and hydrophilic side chains is also predicted by molecular dynamics simulations. The neutron scattering curves of aqueous solutions of hydrophobic amino acids at room temperature are qualitatively similar to differences between the liquid molecular structure functions measured for ambient and supercooled water. The nonpolar solute-induced expansion of water structure reported here is also complementary to recent neutron experiments where compression of aqueous solvent structure has been observed at high salt concentration.

Chemical complementation identifies a proton acceptor for redox-active tyrosine D in photosystem II

Through the use of site-directed mutagenesis and chemical rescue, we have identified the proton acceptor for redox-active tyrosine D in photosystem II (PSII). Effects of chemical rescue on the tyrosyl radical were monitored by EPR spectroscopy. We also have acquired the Fourier–transform infrared (FT-IR) spectrum associated with the oxidation of tyrosine D and concomitant protonation of the acceptor. Mutant and isotopically labeled PSII samples are used to assign vibrational lines in the 3,600–3,100 cm−1 region to N-H modes of His-189 in the D2 polypeptide. When His-189 in D2 is changed to a leucine (HL189D2) in PSII, dramatic alterations of both EPR and FT-IR spectra are observed. When imidazole is introduced into HL189D2 samples, results from both EPR and FT-IR spectroscopy argue that imidazole is functionally reconstituted into an accessible pocket and that imidazole acts as a chemical mimic for His-189. Small perturbations of EPR and FT-IR spectra are consistent with access to this pocket in wild-type PSII, as well. Structures of the analogous site in bacterial reaction centers suggest that an accessible pocket, large enough to contain imidazole, is bordered by tyrosine D and His-189 in the D2 polypeptide. These data provide evidence that His-189 in the D2 polypeptide of PSII acts as a proton acceptor for redox-active tyrosine D and that proton transfer to the imidazole ring facilitates the efficient oxidation/reduction of tyrosine D.

Experimental evidence for hydrogen-bonded network proton transfer in bacteriorhodopsin shown by Fourier-transform infrared spectroscopy using azide as catalyst.

Experimental evidence for proton transfer via a hydrogen-bonded network in a membrane protein is presented. Bacteriorhodopsin's proton transfer mechanism on the proton uptake pathway between Asp-96 and the Schiff base in the M-to-N transition was determined. The slowdown of this transfer by removal of the proton donor in the Asp-96-->Asn mutant can be accelerated again by addition of small weak acid anions such as azide. Fourier-transform infrared experiments show in the Asp-96-->Asn mutant a transient protonation of azide bound to the protein in the M-to-N transition and, due to the addition of azide, restoration of the IR continuum band changes as seen in wild-type bR during proton pumping. The continuum band changes indicate fast proton transfer on the uptake pathway in a hydrogen-bonded network for wild-type bR and the Asp-96-->Asn mutant with azide. Since azide is able to catalyze proton transfer steps also in several kinetically defective bR mutants and in other membrane proteins, our finding might point to a general element of proton transfer mechanisms in proteins.

Femtosecond observation of benzyne intermediates in a molecular beam: Bergman rearrangement in the isolated molecule

In this communication, we report our femtosecond real-time observation of the dynamics for the three didehydrobenzene molecules (p-, m-, and o-benzyne) generated from 1,4-, 1,3-, and 1,2-dibromobenzene, respectively, in a molecular beam, by using femtosecond time-resolved mass spectrometry. The time required for the first and the second C-Br bond breakage is less than 100 fs; the benzyne molecules are produced within 100 fs and then decay with a lifetime of 400 ps or more. Density functional theory and high-level ab initio calculations are also reported herein to elucidate the energetics along the reaction path. We discuss the dynamics and possible reaction mechanisms for the disappearance of benzyne intermediates. Our effort focuses on the isolated molecule dynamics of the three isomers on the femtosecond time scale.

Progress toward chemical accuracy in the computer simulation of condensed phase reactions.

We describe a procedure for the generation of chemically accurate computer-simulation models to study chemical reactions in the condensed phase. The process involves (i) the use of a coupled semiempirical quantum and classical molecular mechanics method to represent solutes and solvent, respectively; (ii) the optimization of semiempirical quantum mechanics (QM) parameters to produce a computationally efficient and chemically accurate QM model; (iii) the calibration of a quantum/classical microsolvation model using ab initio quantum theory; and (iv) the use of statistical mechanical principles and methods to simulate, on massively parallel computers, the thermodynamic properties of chemical reactions in aqueous solution. The utility of this process is demonstrated by the calculation of the enthalpy of reaction in vacuum and free energy change in aqueous solution for a proton transfer involving methanol, methoxide, imidazole, and imidazolium, which are functional groups involved with proton transfers in many biochemical systems. An optimized semiempirical QM model is produced, which results in the calculation of heats of formation of the above chemical species to within 1.0 kcal/mol (1 kcal = 4.18 kJ) of experimental values. The use of the calibrated QM and microsolvation QM/MM (molecular mechanics) models for the simulation of a proton transfer in aqueous solution gives a calculated free energy that is within 1.0 kcal/mol (12.2 calculated vs. 12.8 experimental) of a value estimated from experimental pKa values of the reacting species.