Large terminase conformational change induced by connector binding in bacteriophage T7.

During bacteriophage morphogenesis DNA is translocated into a preformed prohead by the complex formed by the portal protein, or connector, plus the terminase, which are located at an especial prohead vertex. The terminase is a powerful motor that converts ATP hydrolysis into mechanical movement of the DNA. Here, we have determined the structure of the T7 large terminase by electron microscopy. The five terminase subunits assemble in a toroid that encloses a channel wide enough to accommodate dsDNA. The structure of the complete connector-terminase complex is also reported, revealing the coupling between the terminase and the connector forming a continuous channel. The structure of the terminase assembled into the complex showed a different conformation when compared with the isolated terminase pentamer. To understand in molecular terms the terminase morphological change, we generated the terminase atomic model based on the crystallographic structure of its phage T4 counterpart. The docking of the threaded model in both terminase conformations showed that the transition between the two states can be achieved by rigid body subunit rotation in the pentameric assembly. The existence of two terminase conformations and its possible relation to the sequential DNA translocation may shed light into the molecular bases of the packaging mechanism of bacteriophage T7.

Hydration of barium monohydroxide in (H2O)(1-3) clusters: theory and experiment.

The ionization energies (IEe's) of small BaOH(H2O)m clusters (m = 1-3), as generated in a laser vaporization-supersonic expansion source have been determined by laser photoionization experiments over the 3.65-4.55 eV energy range. Complementary ab initio studies show that the IEe's are in good agreement with computed adiabatic ionization energies and that BaOH(H2O)m structures with a direct coordination of the Ba atom to water molecules are favored over those that are characterized by H-bonded networks involving H2O molecules and the OH group of BaOH. Additional calculations have been performed on the hydration energies for the most stable isomers of the relevant BaOH(H2O)1-3 clusters. A comparison is made between the closed-shell title system and the results of related theoretical studies on the open-shell alkali monohydroxides, which allows for an interpretation of the opposite trends that are found in the cluster size dependence of the vertical ionization energies for both series of systems, and highlights the role of the BaOH unpaired electron in its ionization process. Altogether, the present evidence suggests for the initial steps of the BaOH hydration process to be dominated by electrostatic and polarization interactions between the Ba(+) and OH(-) ion cores, which become both increasingly solvated upon sequential addition of water molecules.

Photoionization and ab initio study of Ba(H2O)n (n = 1-4) clusters.

An experimental and theoretical study of the photoionization energies (IE's) of Ba(H(2)O)(n) clusters containing up to n = 4 water molecules has been performed. The clusters were generated by a pick-up source combining laser vaporization with pulsed supersonic expansion, and then photoionized by radiation of 272.5-340 nm. The experimentally determined IE(e)'s for n = 1 to 4 are 4.56 ± 0.05, 4.26 ± 0.05, 3.90 ± 0.05 and 3.71 ± 0.05 eV. This cluster size dependence of IE is reproduced within ±0.06 eV employing the mPW1PW91 density-functional and CCSD(T, Full) quantum-chemical methods combined with the 6-311++G(d,p) basis set for the H and O atoms and three different relativistic effective core potentials for Ba atoms. The calculations indicate that the lowest energy hydration structures represent the most relevant contributions to both the vertical and adiabatic ionization energies. Experimental and theoretical evidence correlates with the progressive surface-delocalization of the electron from the hydration cavity around the Ba atom and suggests that the intra-cluster electron transfer is possible even for small aggregates.

The role of orbiting resonances in the vibrational relaxation of I(2)(B,v' = 21) by collisions with He at very low energies: a theoretical and experimental study.

The low-energy collisions of I(2)(B,v' = 21) with He involving collision-induced vibrational relaxation of I(2) are investigated both experimentally and by means of wave packet simulations. The theoretical cross sections exhibit a structure of peaks originated by orbiting resonances of the I(2)(B,v' = 21) - He van der Waals complex formed in the I(2) + He collisions. Such a structure has similar characteristics as the structure of peaks found in the experimental cross sections. In fact, four of the five peaks of the measured cross sections appear at positions nearly coincident with those of four of the peaks found in the theoretical cross sections. Thus this result confirms the experimental finding that enhancement of I(2) vibrational relaxation is caused by the population of I(2)(B,v' = 21) - He orbiting resonances populated upon the low-energy collisions. The possibility of using this mechanism in the vibrational cooling of diatomic molecules is discussed.

H-bonded network rearrangements in the S0, S1 and D0 states of neutral and cationic p-cresol(H2O)(NH3) complexes.

The H-bonded network rearrangements in the S(0), S(1) and D(0) states of the neutral and cationic p-CreOH(H(2)O)(NH(3)) complexes were studied experimentally by means of (1 + 1)/(1 + 1') REMPI (Resonantly Enhanced MultiPhoton Ionization) and time resolved LIF (Laser Induced Fluorescence) spectroscopies combined with DFT (Density Functional Theory) calculations at the B3LYP/6-311G++(d,p) level. A comparison of the rearrangement process of the H-bonded network in the three states is given. Two cyclic H-bonded isomers were found on the S(0) potential energy surface and the results indicate that the rearrangement in this state is unlikely at the temperature of the supersonic expansion due to the presence of a high-energy barrier (7503 cm(-1)). On the other hand, the re-determination of the S(1) excited state lifetimes confirms that neither the H-bonded rearrangement nor the excited state hydrogen transfer (ESHT) reaction takes place in the S(1) state at the excitation energies of this work. Thus, it is concluded that the absorption of the second photon to reach the D(0) state takes place from the S(1) state of the cyclic-(OH-OH(2)-NH(3)) isomer. A preferential evaporation of H(2)O upon vertical ionization of the cyclic-(OH-OH(2)-NH(3)) isomer is observed which is consistent with a statistical redistribution of the internal energy. Nevertheless, our theoretical calculations suggest that initial excitation of the H-bonded network rearrangement modes may also play a role to leave the H(2)O molecule as a terminal moiety in a chain-(OH-NH(3)-OH(2))(+) isomer. The reaction pathway for the solvent rearrangement involves a double proton transfer process with a very low energy barrier (575 cm(-1)) that is overcome at the vertical ionization energy of the complex.

New determination of the adiabatic ionization potential of the BaOH radical from laser photoionization-molecular beam experiments and ab initio calculations.

The adiabatic ionization potential of the BaOH radical, as generated in a laser vaporization-supersonic expansion source has been determined by laser photoionization experiments to be (4.55 ± 0.03) eV. This value supports the three lowest out of seven previous experimental estimates, the former ranging from 4.35 to 4.62 eV. The present result is compared to ab initio calculations, as performed using both quantum chemistry at different levels of theory and density functional theory, and trying several effective core potentials and their accompanying basis sets for Ba. The most satisfactory agreement is obtained for either the adiabatic or vertical ionization potentials that derive from post-Hartree-Fock [MP2 and CCSD(T)] treatments of electron correlation, along with consideration of relativistic effects and extensive basis sets for Ba, in both BaOH and BaOH(+). Such conclusions extend to the results of related calculations on the Ba-OH dissociation energies of BaOH and BaOH(+), which were performed to help in calibrating the present computational study. Bonding in BaOH/BaOH(+), as well as possible sources of discrepancy with previous experimental determinations of the BaOH adiabatic ionization potential are discussed.

Unexpected size distribution of Ba(H2O)n clusters: why is the intensity of the Ba(H2O)1 cluster anomalously low?

An experimental and theoretical study on the reactivity of neutral Ba atoms with water clusters has been conducted to unravel the origin of the irregular intensity pattern observed in one-photon ionization mass spectra of a Ba(H(2)O)(n)/BaOH(H(2)O)(n-1) (n = 1-4) cluster distribution, which was generated in a laser vaporization-supersonic expansion source. The most remarkable irregular feature is the finding for n = 1 of a lower intensity for the Ba(+)(H(2)O)(n) peak with respect to that of BaOH(+)(H(2)O)(n-1), which is opposite to the trend for n = 2-4. Rationalization of the data required consideration of a distinct behavior of ground-state and electronically excited state Ba atoms in inelastic and reactive Ba + (H(2)O)(n) encounters that can occur in the cluster source. Within this picture, the generation of Ba(H(2)O)(n) (n > 1) association products results from stabilizing collisions with atoms of the carrier gas, which are favored by intramolecular vibrational redistribution that operates on the corresponding collision intermediates prior to stabilization; the latter is unlikely to occur for Ba + (H(2)O) encounters. Overall, this interpretation is consistent with additional in-source laser excitation and quenching experiments, which aimed to explore qualitatively the effect of perturbing the Ba atom electronic state population distribution on the observed intensity pattern, as well as with the energetics of various possible reactions for the Ba + H(2)O system that derive from high level ab initio calculations.

Chemiluminescence from the Ba((3)P)+N(2)O-->BaO(A (1)Sigma(+))+N(2) reaction: Collision energy effects on the product rotational alignment and energy release.

Both fully dispersed unpolarized and polarized chemiluminescence spectra from the Ba((3)P)+N(2)O reaction have been recorded under hyperthermal laser-ablated atomic beam-Maxwellian gas conditions at three specific average collision energies E(c) in the range of 4.82-7.47 eV. A comprehensive analysis of the whole data series suggests that the A (1)Sigma(+)-->X (1)Sigma(+) band system dominates the chemiluminescence. The polarization results revealed that the BaO(A (1)Sigma(+)) product rotational alignment is insensitive to its vibrational state upsilon(') at E(c)=4.82 eV but develops into an strong negative correlation between product rotational alignment and upsilon(') at 7.47 eV. The results are interpreted in terms of a direct mechanism involving a short-range, partial electron transfer from Ba((3)P) to N(2)O which is constrained by the duration of the collision, so that the reaction has a larger probability to occur when the collision time is larger than the time needed for N(2)O bending. The latter in turn determines that, at any given E(c), collinear reactive intermediates are preferentially involved when the highest velocity components of the corresponding collision energy distributions are sampled. Moreover, the data at 4.82 eV suggest that a potential barrier to reaction which favors charge transfer to bent N(2)O at chiefly coplanar geometries is operative for most of the reactive trajectories that sample the lowest velocity components. Such a barrier would arise from the relevant ionic-covalent curve crossings occurring in the repulsive region of the covalent potential Ba((3)P)cdots, three dots, centeredN(2)O((1)Sigma(+)); from this crossing the BaO(A (1)Sigma(+)) product may be reached through mixings in the exit channel with potential energy surfaces leading most likely to the spin-allowed b (3)Pi and a (3)Sigma(+) products. The variation with increasing E(c) of both the magnitude of the average BaO(A (1)Sigma(+)) rotational alignment and the BaO(A (1)Sigma(+)) rovibrational excitation, as obtained from spectral simulations of the unpolarized chemiluminescence spectra, consistently points to additional dynamic factors, most likely the development of induced repulsive energy release as the major responsible for the angular momentum and energy disposal at the two higher E(c) studied. The results of a simplified version of the direct interaction with product repulsion-distributed as in photodissociation model do not agree with the observed average product rotational alignments, showing that a more realistic potential energy surface model will be necessary to explain the present results.

Effect of the intermolecular hydrogen bond conformation on the structure and reactivity of the p-cresol(H2O)(NH3) van der Waals complex.

The structure and reactivity of p-CrOH(NH(3))(2) and p-CrOH(H(2)O)(NH(3)) complexes were studied using mass-resolved one-colour resonance-enhanced multi-photon ionization and laser-induced fluorescence (LIF) spectroscopy together with DFT calculations. At the excitation energy of this work, the S(1) state of p-CrOH(NH(3))(2) shows a sub-nanosecond lifetime, as determined by time-resolved LIF spectra, as a consequence of a hydrogen transfer process that results in NH(4)(NH(3)) as a reaction product. Substitution of NH(3) by H(2)O closes the reaction channel as evidenced by the absence of excited-state hydrogen transfer (ESHT) reaction products, (H(3)O(NH(3)) or NH(4)(H(2)O)) and results in a dramatic effect on the S(1) lifetime of the p-CrOH(H(2)O)(NH(3)) complex which rises to (12 +/- 2) ns. According to density functional theory calculations, the most stable isomer of the p-CrOH(H(2)O)(NH(3)) complex is a cyclic structure, in which H(2)O acts as the H acceptor of the phenolic OH group (c-OH-H(2)O-NH(3)). However, the ESHT process is energetically disallowed upon electronic excitation.

Vibrational energy transfer between I2(B (3)Pi(0u)+, nu' = 21) and He at very low temperatures: impulsive versus complex formation mechanisms assisted by tunneling through the centrifugal barrier.

The temperature dependence of the state-to-state vibrational relaxation rate constant (k(nu)(21-Delta nu)) for collisions between I(2)(B,nu(')=21) and He at very low kinetic energies was studied. The fluorescence from I(2)(B,nu(')=21-Delta nu(')) with Delta nu(')=1-5 indicates that in the temperature range of 0.6-8.2 K these states are populated by only one collision with He. The behavior of k(nu)(21-Delta nu) with temperature can be divided into two groups. The group with quantum changes Delta nu(')=1-3 shows scattering resonances in the low temperature region, with a general monotonical decrease of the rate constant with temperature, suggesting the importance of van der Waals interactions. This behavior is supported by the calculation of the probability of tunneling through the centrifugal barriers. For collisions in which 4-5 quanta are lost in a single event, there are no evidences of scattering resonances and the values of the relaxation rate constants could be determined only at the highest temperatures of this study. This suggests that relaxation occurs via impulsive collisions. The branching ratios for each channel are also temperature dependent and this behavior also suggests that the energy transfer mechanism changes with Delta nu(').

Mutational analysis of the pH signal transduction component PalC of Aspergillus nidulans supports distant similarity to BRO1 domain family members.

The alkaline ambient pH signal transduction pathway component PalC has no assigned molecular role. Therefore we attempted a gene-specific mutational analysis and obtained 55 new palC loss-of-function alleles including 24 single residue substitutions. Refined similarity searches reveal conserved PalC regions including one with convincing similarity to the BRO1 domain, denoted PCBROH, where clustering of mutational changes, including PCBROH key residue substitutions, supports its structural and/or functional importance. Since the BRO1 domain occurs in the multivesicular body (MVB) pathway protein Bro1/Vps31 and also the pH signal transduction protein PalA (Rim20), both of which interact with MVB component (ESCRT-III protein) Vps32/Snf7, this might reflect a further link between the pH response and endocytosis.

Building transition probabilities for any condition using reduced cumulative energy transfer functions in H2O-H2O collisions.

The energy transfer process between highly vibrationally excited H(2)O in thermal equilibrium with a gas bath of H(2)O at different internal energies and temperatures has been studied by classical trajectory calculations. The results were analyzed using a cumulative probability distribution Q(DeltaE) of the amount of energy transferred, obtained by direct count of the number of trajectories that transfer an amount of energy equal to or greater than a certain value DeltaE. Scaling Q(DeltaE) in terms of the mean down and up energies transferred for each group of trajectories results in a unique distribution. This fact and the use of detailed balance constrains were used to propose a methodology that make it possible to build the whole P(E('),E) for any condition by knowing DeltaE and a series of parameters that depend only on the system under study.

Accounting for the dependence of P(E',E) on the maximum impact parameter in classical trajectory calculations: application to the H2O-H2O collisional relaxation.

In this work we report a novel methodology that is able to predict how energy transfer transition probability density functions [P(E',E)] change with the maximum impact parameter (bmax) used in trajectory calculations (TC's). The method assumes that P(E',E) can be described by a sum of exponential functions and that all the trajectories with an initial impact parameter beyond a certain critical value will contribute only to the elastic peak [P(E',E) for E'=E]. This approach is applied to H2O-H2O collisions at different initial vibrational energies of the excited molecules and temperatures of bath gas. The results show that it is possible to reproduce with high accuracy the whole P(E',E) obtained from a given bmax, using the results of TC's performed at another bmax. The new methodology also leads us to propose a new criterion to choose the value of bmax.

Chemiluminescent reaction of Ba(3P) with N2O at hyperthermal collision energies: rotational alignment of the BaO(A 1Sigma+) product.

The chemiluminescent reaction Ba(6s6p (3)P)+N(2)O was studied at an average collision energy of 1.56 eV in a beam-gas arrangement. Ba((3)P) was produced by laser ablation of barium, which resulted in a broad collision energy distribution extending up to approximately 5.7 eV. A series of experiments was made to extract the Ba((3)P) contribution to chemiluminescence from that corresponding to Ba 6s(2) (1)S0 and 6s5d (3)D, which are the other two most populated states in the atomic beam. The fully dispersed polarized chemiluminescence spectra at 400-600 nm from the title reaction were recorded and assigned to a BaO molecule excited in the A (1)Sigma+ level. In addition, the average and wavelength-resolved degrees of polarization associated to the parallel BaO(A (1)Sigma+-->X (1)Sigma+) emission are reported. The analysis of the average polarization degree show that the BaO(A (1)Sigma+) product is significantly aligned, suggesting that the reaction mechanism is predominantly direct. The product rotational alignment was found to depend markedly on the emission wavelength, which revealed a negative correlation with the BaO(A (1)Sigma+) product vibrational state. On the basis of experimental and theoretical investigations on the reactions of N(2)O with both the (1)S0, (3)D, and (1)P1 states of Ba and the lighter group 2 atoms, it is suggested that the Ba((3)P) reaction involves a charge transfer at relatively short reagent separations and that restricted collision geometries at the highest velocity components of the broad distribution are necessary to rationalize the data.

Analysis of arsenic and calcium in soil samples by laser ablation mass spectrometry.

We present an analytical procedure based on laser ablation mass spectrometry (LAMS) in order to detect and quantify arsenic and calcium in soil samples and we analyze the diverse factors that influence the precision of LAMS, such as laser fluence and matrix effect. The results indicate that a Zn matrix is a good choice for the analysis of those metals in soil samples. This work also provides a method for the direct determination of As in soil samples whose concentrations are lower than 100 ppm with a 70 ppm minimum detection limits (MDL).

Fitting complex potential energy surfaces to simple model potentials: application of the simplex-annealing method.

A stochastic method of optimization, which combines simulated annealing with simplex, is implemented to fit the parameters of a simple model potential. The main characteristic of the method is that it explores the whole space of the parameters of the model potential, and therefore it is very efficient in locating the global minimum of the cost function, in addition to being independent of the initial guess of the parameters. The method is employed to fit the complex intermolecular potential energy surface of the dimer of water, using as a reference the spectroscopic quality anisotropic site-site potential of Feller et al. The simple model potential chosen for its reparameterization is the MCY model potential of Clementi et al. The quality of the fit is assessed by comparing the geometry of the minimum, the harmonic frequencies, and the second virial coefficients of the parameterized potential with the reference one. Finally, to prove more rigorously the robustness of this method, it is compared with standard nonstochastic methods of optimization.

Arrestin-related proteins mediate pH signaling in fungi.

Metazoan arrestins bind to seven-transmembrane (7TM) receptors to regulate function. Aspergillus nidulans PalF, a protein involved in the fungal ambient pH signaling pathway, contains arrestin N-terminal and C-terminal domains and binds strongly to two different regions within the C-terminal cytoplasmic tail of the 7TM, putative pH sensor PalH. Upon exposure to alkaline ambient pH, PalF is phosphorylated and, like mammalian beta-arrestins, ubiquitinated in a signal-dependent and 7TM protein-dependent manner. Substitution in PalF of a highly conserved arrestin N-terminal domain Ser residue prevents PalF-PalH interaction and pH signaling in vivo. Thus, PalF is the first experimentally documented fungal arrestin-related protein, dispelling the notion that arrestins are restricted to animal proteomes. Epistasis analyses demonstrate that PalF posttranslational modification is partially dependent on the 4TM protein PalI but independent of the remaining pH signal transduction pathway proteins PalA, PalB, and PalC, yielding experimental evidence bearing on the order of participation of the six components of the pH signal transduction pathway. Our data strongly implicate PalH as an ambient pH sensor, possibly with the cooperation of PalI.