Electrostatic Contributions to Protein Stability and Folding Energy

The ability to predict the thermal stability of proteins based on their corresponding sequence is a problem of great fundamental and practical importance. Here we report an approach for calculating the electrostatic contribution to protein stability based on the use of the semimacroscopic protein dipole Langevin dipole (PDLD/S) in its linear response approximation version for self-energy with a dielectric constant, (εpεp) and an effective dielectric for charge–charge interactions (εeffεeff). The method is applied to the test cases of ubiquitin, lipase, dihydrofolate reductase and cold shock proteins with series of εpεp and εeffεeff. It is found that the optimal values of these dielectric constants lead to very promising results, both for the relative stability and the absolute folding energy. Consideration of the specific values of the optimal dielectric constants leads to an exciting conceptual description of the reorganization effect during the folding process. Although this description should be examined by further microscopic studies, the practical use of the current approach seems to offer a powerful tool for protein design and for studies of the energetics of protein folding.This work was supported by NIH Grant GM2449

Substrate promiscuity in DNA methyltransferase M.PvuII. A mechanistic insight

M.PvuII is a DNA methyltransferase from the bacterium Proteus vulgaris that catalyzes methylation of cytosine at the N4 position. This enzyme also displays promiscuous activity catalyzing methylation of adenine at the N6 position. In this work we use QM/MM methods to investigate the reaction mechanism of this promiscuous activity. We found that N6 methylation in M.PvuII takes place by means of a stepwise mechanism in which deprotonation of the exocyclic amino group is followed by the methyl transfer. Deprotonation involves two residues of the active site, Ser53 and Asp96, while methylation takes place directly from the AdoMet cofactor to the target nitrogen atom. The same reaction mechanism was described for cytosine methylation in the same enzyme, while the reversal timing, that is methylation followed by deprotonation, has been described in M.TaqI, an enzyme that catalyzes the N6-adenine DNA methylation from Thermus aquaticus. These mechanistic findings can be useful to understand the evolutionary paths followed by N-methyltransferases.This work was supported by the Ministerio de Ciencia e Innovación project CTQ2009-14541-C02-02, Generalitat Valenciana projects ACOMP/2011/028, ACOM/2012/243, GV/2012/053 and Universitat de Valencia project UV-INV-AE11-40931. J.A. and M.R. thank Ministerio Ciencia e Innovación for a doctoral grant and a ‘Juan de la Cierva’ contract, respectively. The authors acknowledge computational facilities of the Servei d’Informàtica de la Universitat de València in the ‘Tirant’ supercomputer, which is part of the Spanish Supercomputing Network

Critical evaluation of anharmonicity and configurational averaging in QM/MM modelling of equilibrium isotope effects

Anharmonic effects upon vibrational frequencies and isotopic partition function ratios are modelled computationally by means of quantum mechanics/molecular mechanics (QM/MM) methods for two systems. First, the methyl cation in explicit water is considered using a B3LYP/6-31+G(d)/TIP3P method in order to check the previous prediction of an inverse equilibrium isotope effect (EIE) KH3/KD3 for transfer from vacuum to water at 298 K. A full QM/MM treatment including Lennard-Jones interactions predicts significantly inverse contributions from both internal (0.843 0.001) and external (0.894 0.001) modes of the solute. This treatment yields a much larger harmonic EIE (0.753 0.002, averaged over 928 independent solvent configurations) than is obtained either by projecting out the translational and rotational contributions (0.853) or by treating the solvent by a point-charge representation (0.9360 0.0006, harmonic; 0.9366 0.0006, anharmonic). The contribution of anharmonicity to the EIE affects the value only in the 3rd significant figure. Second, anharmonicity is investigated by means of QM/MM potential-energy scans along 12 normal modes for internal and external vibrations of methyl cation in water and for three modes (one stretching and two bending) for the Ha atom at the carbenium-ion centre in cyclopentyl, cyclohexyl, tetrahydrofuranyl and tetrahydropyranyl cations in explicit water and cyclohexane solvents, as obtained by means of atomic Hessian analysi

The Osireion of Oxyrhynchus (El Bahnasa, Egipto)

En el año 2000 se descubrió, al oeste de la ciudad grecorromana de Oxirrinco, una serie de estructuras subterráneas consagradas a Osiris. Bajo una elevación natural del terreno se ha iniciado la excavación de un recinto cultual presidido por una estatua colosal del dios y, adyacente a esta sala, un pasillo donde se enterraban los simulacros de Osiris que se realizaban con motivo del ritual de los misterios del mes de Khoiak. En las dos últimas campañas los trabajos se han centrado en la superestructura del monumento. Se ha localizado el témenos, pudiendo, así, delimitar toda el área del recinto sagrado. Se ha excavado, también, la entrada monumental por la que se accedía al interior de la catacumba. Delante de la entrada, una serie de estructuras nos revelan una zona donde se realizaban las ofrendas diarias a la divinidad. Hasta el momento, podemos fechar el conjunto en época ptolemaicaIn the year 2000, a series of underground structures dedicated to Osiris were discovered to the west of the Graeco-Roman city of Oxyrhynchus. Beneath a natural slope in the land, excavation was started of an enclosure used for worship, presided over by a colossal statue of the god. Next to this chamber was a passage where the figurines of Osiris that were made to be used in the ritual mysteries of the month of Khoiak were buried. In the last two campaigns, work has concentrated on the superstructure of the monument. The temenos had been located thus allowing the limits of the area used for worship to be delineated. The monumental entrance which gave access to the interior of the catacomb has also been excavated. In front of the entrance there is a series of structures that reveal an area where daily offerings were made to the divinity. So far we can date the complex to the Ptolemaic period

On the relationship between thermal stability and catalytic power of enzymes

The  possible  relationship  between  the  thermal  stability  and  the  catalytic  power  of  enzymes  is  of   great  current  interest.  In  particular,  it  has  been  suggested  that  thermophilic  or  hyperthermophilic   (Tm)   enzymes   have   lower   catalytic   power   at   a   given   temperature   than   the   corresponding   mesophilic   (Ms)   enzymes,   because   the   thermophilic   enzymes   are   less   flexible   (assuming   that   flexibility   and   catalysis   are   directly   correlated).   These   suggestions   presume   that   the   reduced   dynamics   of   the   thermophilic   enzymes   is   the   reason   for   their   reduced   catalytic   power.   The   present  paper  takes  the  specific  case  of  dihydrofolate  reductase  (DHFR) and explores the validity of the above argument by simulation approaches. It is found that the Tm enzymes have restricted motions in the direction of the folding coordinate, but this is not relevant to the chemical process, since the motions along the reaction coordinate are perpendicular to the folding motions. Moreover, it is shown that the rate of the chemical reaction is determined by the activation barrier and the corresponding reorganization energy, rather than by dynamics or flexibility in the ground state. In fact, as far as flexibility is concerned, we conclude that the displacement along the reaction coordinate is larger in the Tm enzyme than in the Ms enzyme and that the general trend in enzyme catalysis is that the best catalyst involves less motion during the reaction than the less optimal catalyst. The relationship between thermal stability and catalysis appears to reflect the fact that in order to obtain small electrostatic reorganization energy it is necessary to invest some folding energy in the overall preorganization process. Thus, the optimized catalysts are less stable. This trend is clearly observed in the DHFR case

Reversibility and Diffusion in Mandelythiamin Decarboxylation. Searching Dynamical Effects in Decarboxylation Reactions

Decarboxylation of mandelylthiamin in aqueous solution is analyzed by means of quantum mechanics/molecular mechanics simulations including solvent effects. The free energy profile for the decarboxylation reaction was traced, assuming equilibrium solvation, while reaction trajectories allowed us to incorporate nonequilibrium effects due to the solvent degrees of freedom as well as to evaluate the rate of the diffusion process in competition with the backward reaction. Our calculations that reproduce the experimental rate constant show that decarboxylation takes place with a non-negligible free energy barrier for the backward reaction and that diffusion of carbon dioxide is very fast compared to the chemical step. According to these findings catalysts would not act by preventing the backward reaction.GV/2012/053. ESTUDIO DEL MECANISMO DE REACCION Y DE LOS MOVIMIENTOS DINAMICOS DE LAS ENZIMAS ADN-METILTRANSFERASAS: CATALISIS Y INHIBICIO

Fourier analysis of the aerodynamic behavior of cup anemometers

The calibration results (the transfer function) of an anemometer equipped with several cup rotors were analyzed and correlated with the aerodynamic forces measured on the isolated cups in a wind tunnel. The correlation was based on a Fourier analysis of the normal-to-the-cup aerodynamic force. Three different cup shapes were studied: typical conical cups, elliptical cups and porous cups (conical-truncated shape). Results indicated a good correlation between the anemometer factor, K, and the ratio between the first two coefficients in the Fourier series decomposition of the normal-to-the-cup aerodynamic forc

Fabrication of glass-based products as remediation alternative for contaminated urban soils of Barcelona

Peer ReviewedObjectius de Desenvolupament Sostenible::12 - Producció i Consum ResponsablesPostprint (published version

Big Eight +12. : Test de competències d'empleabilitat

Test Situacional de Competències d'Empleabilitat per a joves de 12 a 18 anysEls resultats que s'obtenen en aquest test serveixen per reflexionar sobre el desenvolupament de les competències d'empleabilitat. Aquestes competències no són un tret de la personalitat, sinó que són resultat de l'aprenentatge i les experiències vitals i, per tant, subjectes de desenvolupament i de millora

Temperature dependence of dynamic, tunnelling and kinetic isotope effects in formate dehydrogenase

The origin of the catalytic power of enzymes has been a question of debate for a long time. In this regard, the possible contribution of protein dynamics in enzymatic catalysis has become one of the most controversial topics. In the present work, the hydride transfer step in the formate dehydrogenase (FDH EC 1.2.1.2) enzyme is studied by means of molecular dynamic (MD) simulations with quantum mechanics/molecular mechanics (QM/MM) potentials in order to explore any correlation between dynamics, tunnelling effects and the rate constant. The temperature dependence of the kinetic isotope effects (KIEs), which is one of the few tests that can be studied by experiments and simulations to shed light on this debate, has been computed and the results have been compared with previous experimental data. The classical mechanical free energy barrier and the number of recrossing trajectories seem to be temperature-independent while the quantum vibrational corrections and the tunnelling effects are slightly temperature-dependent over the interval of 5–45 °C. The computed primary KIEs are in very good agreement with previous experimental data, being almost temperature-independent within the standard deviations. The modest dependence on the temperature is due to just the quantum vibrational correction contribution. These results, together with the analysis of the evolution of the collective variables such as the electrostatic potential or the electric field created by the protein on the key atoms involved in the reaction, confirm that while the protein is well preorganised, some changes take place along the reaction that favour the hydride transfer and the product release. Coordinates defining these movements are, in fact, part of the real reaction coordinate