Identification,Cloning and Biochemical Characterization of Pseudomonas putida A (ATCC 12633) Monooxygenase Enzyme necessary for the Metabolism of Tetradecyltrimethylammonium Bromide

This study presents the first report of the purification and characterization of a monooxygenase enzyme from Pseudomonas putida A (ATCC 12633) that is responsible for the oxidation of physiologically relevant quaternary ammonium compounds, the tetradecyltrimethylammonium bromide. The degradation of tetradecyltrimethylammonium bromide by P. putida A (ATCC 12633) is initiated by N-dealkylation and catalysed by tetradecyltrimethylammonium monooxygenase (TTABMO), resulting in the formation of tetradecylalkanal and trimethylamine. Based on sequence analysis, the gene for TTABMO (ttbmo) corresponded to an ORF named PP2033 in the genome of P. putida KT2440. Mutation in ttabmo blocked the utilization of tetradecyltrimethylammonium bromide by Pseudomonas putida A (ATCC 12633) as carbon and nitrogen sources. The enzyme can be highly overexpressed in P. putida Δttabmo-T7 in active form and purified as a hexahistidine fusion protein. Like the native enzyme, the his-TTABMO was found to be a monomer with molecular mass of 40 kDa, the isoelectric point 7.3, that catalyses the breakdown of tetradecyltrimethylammonium bromide and utilized NADPH and FAD as cofactor. The biochemical properties and the analysis of the respective protein sequence revealed that TTABMO represents a typical flavoprotein monooxygenase, which is member of a flavoprotein family that is distinct from Baeyer–Villiger monooxygenases.     

Laser spectroscopic investigation of isotope shifts in Nd II lines

The isotope shift of 11 optical transitions in Nd II in the spectral range 420–450 nm have been recorded (in all cases but one for all pairs of even Nd isotopes). For all observed transitions the values of the field shifts, the specific mass shifts and Δ∣ψ(0)∣² have been evaluated. Using our new data, combined with data reported in earlier papers, term isotope shifts for all pairs of even Nd isotopes have been determined for 27 energy levels and for the configurations: 4f⁴6s, 4f⁴5d, 4f³5d6s and 4f⁴6p as well. It is shown that configuration assignment of a level on the basis of term shift values only is in some cases not satisfactory (as e.g. of the level at 22,696 cm⁻¹) and obviously additional information is required.     

Reevaluation of electronic polarization energies in organic molecular crystals

Effective electronic polarization energies for positive P⁺_eff and negative P⁻_eff charge carriers in polyacene crystals have been reevaluated. By comparing the P⁺_eff and P⁻_eff values obtained from the analyses of reported energy parameters with calculated data, it is shown that the widely accepted assumption that P⁺_eff = P⁻_eff in polyacene crystals is not valid. By applying recently reported data on optical E^opt_G and adiabatic E^ad_G energy gap values, the contribution of molecular (vibronic) polarization in the total effective polarization energies W⁺_eff and W⁻_eff has been estimated, and modified energy diagrams for polyacene crystals have been presented. Further, due to practically constant values of observed and calculated P⁺_eff and P⁻_eff in polyacenes, the gap energies between positive and negative charge carrier conduction levels have been estimated for several related aromatic hydrocarbons.     

Evaluation of dsDNA as a wire for redox-active proteins

The redox-active multiligand-binding flavoprotein dodecin binds flavins with high affinity when they are oxidized, whereas flavin reduction induces the dissociation of the holoprotein complex in apododecin and free flavin ligands. Dodecin could be reconstituted at flavin-terminated dsDNA monolayers. The binding and release of apododecin triggered by the redox state of the flavins can be monitored by surface-sensitive techniques such as surface plasmon resonance and quartz crystal microbalance measurements with dissipation monitoring. It has been shown that flavin reduction followed by the release of apododecin can be achieved by mediated electron transfer in the presence of the redox mediator amino ethyl viologen and by chemical flavin reduction, whereas flavin reduction by direct electron transfer via the dsDNA tethers is not possible. The combination of electrochemistry with surface-sensitive techniques such as surface plasmon resonance or quartz crystal microbalance measurements with dissipation monitoring could be highly beneficial to confirm or disprove the mechanism, which has been postulated for the action of primases, which contain a [4Fe4S] cluster and are involved in DNA replication. It has been postulated that these enzymes bind the DNA template when the cluster is in the [4Fe4S]³⁺ state, whereas they are released when the cluster is reduced via electron transfer through DNA and the protein environment.     

Lifetime of 2s2p 23s 3 P in N II and other allowed triplet transitions

Systematic transition calculations have been performed for all triplet decay channels from 2s2p ²3s ³ P of N II. The lifetime of 0.505 ns compares favourably with experiment. Transitions in absorption from the 2s ²2p ^{2 3} P ground state have also been determined along with a number of other triplet transitions. Length and velocity forms of thef-value are in agreement at the 2–3% level for most transitions.     

Hyperfine structure in the configuration 4f 115d6s 2 of Er I

Hyperfine constants of low lying levels of the configuration 4f ¹¹5d6s ² in Er I have been measured for the only stable odd isotope,¹⁶⁷Er, using high resolution laseratomic-beam spectroscopy. Hyperfine parameters were evaluated from the experimental data using the effective tensor operator formalism. For this purpose eigenfunctions have been determined from an analysis of the fine structure energies of Er I as well as from ab initio multiconfiguration Dirac-Fock calculations. With the latter method also ab initio hyperfine constants for the levels investigated were evaluated. A comparison of calculated fine structure energies, Landég _J -factors and hyperfine constants with the experimental data allowed a test of the reliability of the fine structure and multiconfiguration Dirac-Fock wavefunctions. Effective nuclear electric quadrupole moments for¹⁶⁷Er have been determined from the experimental hyperfine constants using both calculated relativistic electronic radial integrals and hyperfine parameters for the 4f and 5d electrons in the configuration 4f ¹¹5d6s ² in Er I. From a comparison with the nuclear quadrupole moment measured in the mesic atom Sternheimer shielding factors are calculated. Configuration-interaction contributions to the radial integrals 〈r ^?3〉 _nl ⁰¹ of the 4f and 5d electrons have been determined.     

Identification of new lines in the beam-foil spectrum of B II–IV

Ten, six, and twenty-one new lines appearing in the beam-foil spectra of boron (30–200 nm) have been identified in B II, B III, and B IV, respectively. Energies have been determined with higher accuracy than obtained before for the 2p3s ¹ P ⁰, 2s6d ³ D, 2p3p ³ D, 2p3p ³ S, 2p3p ³ P, and 2p3p ¹ D terms in B II. First energy values are also reported for the 2s6d ¹ D and 2s7d ³ D terms in B II.     

Thermodynamics of molecular interactions in binary mixtures of non-electrolytes. Molar excess volumes

Molar excess volumes, V^E, for pyridine (A) + α-picoline (B), + β-picoline (B) and + γ-picoline (B) and benzene (A) + toluene (B), + o-xylene (B) and + p-xylene (B) and carbon tetrachloride (A) + n-heptane (B) have been measured dilatometrically as a function of temperature and composition and have been utilized to study B—B and B—B—B interactions in the presence of A via the Mayer—McMillan approach. A model has also been presented to account for these B—B and B—B—B interactions. The V^E data at 308.15 K have also been analysed in terms of the “graph theoretical” approach which describes the V^E data well for all these mixtures at 308.15 K. The “graph theoretical” approach has further been extended to successfully evaluate V^E data for a mixture at any temperature, T₂, when the V^E data at T₁ are known.     

Alignment and orientation of Ar+ in He+-Ar collisions

We have measured the alignment and orientation parameters of the² F ₇ ^2/0 and² F ₅ ^2/0 states of Ar⁺ formed in the two-electron process; He⁺+Ar→He(1s ²)+Ar⁺(3p ⁴4p′). These have been measured at a collision energy of 0.25 keV/amu and for scattering angles ranging from 0.94° to 3.75°. First, by comparing the orientation parameter for the Ar⁺[(3p ⁴[¹ D]4p′² F ₇ ^2/0 ] and the Ar⁺[(3p ⁴[¹ D]4p′² F ₅ ^2/0 ] states, we have experimentally determined the importance of the spin-dependent interactions for the present collision system, by testing the Percival-Seaton hypothesis of spin independence. If the Percival-Seaton hypothesis holds for this system, the orientation parameter should beJ-independent. Secondly, the magnitude of the orientation parameter can be interpreted as resulting from the collective circulation of the unexcited 3p ⁴ electrons and the excited 4p electron. The direction of this collective circulation is compared to the propensity rule for colliding di-atom systems.     

Classification of 3-3 transitions in neonlike germanium in laser-produced plasma

Classification of 3-3 transitions in neonlike germanium in laser-produced plasma has been made. The spectra have been observed in the range of 185–290 Å using a high resolving power (λ/Δλ ～ 13000) grazing-incidence spectrometer in XUV laser experiment. A total of 21 lines (including five lasing lines) have been classified as transitions between the 2s ²2p ⁵3s, 3p, 3d or 2s2p ⁶3p, 3d and sodiumlike germanium configurations. The identified transitions have been used to derive energy levels of 2s ²2p ⁵3l in neonlike germanium. The experimental results have been compared with theoretical predictions from Dirac-Fock (MCDF) calculations.     

Theoretical study of LiK and LiK+ in adiabatic representation

The potential energy curves have been calculated for the electronic states of the molecule LiK within the range 3 to 300 a.u., of the internuclear distance R. Using an ab initio method, through a semiempirical spin-orbit pseudo-potential for the Li (1s ²) and K (1s ²2s ²2p ⁶3s ²3p ⁶) cores and core valence correlation correction added to the electrostatic Hamiltonian with Gaussian basis sets for both atoms. The core valence effects including core-polarization and core-valence correlation are taken into account by using an l-dependent core-polarization potential. The molecular orbitals have been derived from self-consistent field (SCF) calculation. The spectroscopic constants, dipole moments and vibrational levels of the lowest electronic states of the LiK molecule dissociating into K (4s, 4p, 5s, 3d, and 5p) + Li (2s, 2p, 3s, and 3p) in ^{1, 3}Σ, ^{1, 3}Π, and ^{1, 3}Δ symmetries. Adiabatic results are also reported for ²Σ, ²Π, and ²Δ electronic states of the molecular ion LiK⁺ dissociating into Li (2s, 2p, 3s, and 3p) + K⁺ and Li⁺ + K (4s, 4p, 5s, 3d, and 5p). The comparison of the present results with those available in the literature shows a very good agreement in spectroscopic constants of some lowest states of the LiK and LiK⁺ molecules, especially with the available theoretical works. The existence of numerous avoided crossing between electronic states of ²Σ and ²Π symmetries is related to the charge transfer process between the two ionic systems Li⁺K and LiK⁺.     

Isotope shift and hyperfine structure measurements in titanium I

High accuracy measurements of hyperfine structure due to⁴⁷Ti and⁴⁹Ti in the 3d ² 4s ² a ³ F ₂?3d ² 4s4p z ⁵ D ₁ absorption line at σ=18482.772 cm^?1 have been performed by use of a Doppler-free experiment, where a beam of titanium atoms is crossed by a CW single mode tunable dye laser. They have allowed for the determination of isotope shifts between⁴⁶Ti,⁴⁷Ti,⁴⁸Ti,⁴⁹Ti and⁵⁰Ti. By use of accurate values of mean square nuclear charge radii for the even isotopes, it has been possible to separate mass shifts from field shifts and to determine accurate values for the mean square nuclear charge radii of⁴⁷Ti and⁴⁹Ti. The field shift presents a marked odd-even staggering.     

Two-color excitation of NO in a supersonic free jet. Autoionization of high rydberg states

High Rydberg states of NO above the ionization limit have been measured for the isolated molecule in a supersonic free jet by two-color multiphoton ionization. Three Rydberg series (ns, np and nf) were identified, which appeared by rotational and the vibrational autoionization. The rotational structures of the 13s(υ = 1), 13p(υ = 1) and 12f(υ = 1) states have been analyzed in detail. The fluorescence dip spectra for the intermediate A²Σ⁺(3sσ) state have been measured simultaneously and the cross sections of the one-photon absorption to the high Rydberg states from the A²Σ⁺(υ = 1) state have been determined.     

Wavelengths and transition probabilities in the quartet spectrum of NV

Energies and structures of 1s2snl ⁴ L and 1s2pnl' ⁴ L core-excited levels in lithium-like NV are calculated. Wavelengths, transition probabilities and radiative lifetimes are reported and compared with available experimental data. It is suggested that the 1s2p4d ⁴ P ⁰,⁴ D ⁰ levels are nearly degenerate in NV and have not been resolved experimentally. Several new assignments for observed lines are proposed.     

Isotope shift studies in the spectra of Gadolinium in UV region and term shifts of high even levels of Gd I

Isotope shift Δσ(¹⁵⁶Gd?¹⁶⁰Gd) is reported in 70 spectral lines of neutral gadolinium atom (Gd I) in the 3290- 3920 Å region providing isotope shift data in UV lines of Gd I spectrum for the first time. The measurements were carried out on a photoelectric recording Fabry-Perot Spectrometer using highly enriched isotopic samples of gadolinium. Term isotope shifts ΔT(¹⁵⁶Gd?¹⁶⁰Gd) have been evaluated for 48 high lying even parity energy levels of Gd I using this data; new ΔT values have been obtained for 24 levels. Electronic configurations 4f⁷5d6s6p, 4ff⁷5df²6p and 4f⁸5d6s assigned earlier to these even levels have been either confirmed or configuration mixing pointed out in some of these levels. Probable assignment of 4f⁸5d6s configuration to 8 even levels between 32 930 and 35 500 cm^-1 have been confirmed.     

Experimental and theoretical investigation of radiative lifetimes in neutral gallium

Using laser spectroscopic techniques the natural radiative lifetimes of 4s ² n ₁ s ² S and 4s ² n ₂ d ² D states of neutral gallium have been measured forn ₁ = 6 to 11 andn ₂ = 4 to 9. These states, as well as previously measured4s ² np ² P states, have been investigated theoretically using multi-configuration Hartree-Fock calculations. Oscillator strengths to all lower-lying states have been calculated and theoretical lifetimes of the investigated states evaluated. The² D sequence is strongly influenced by the 4s4p ^{2 2} D perturber, and strong cancellation effects in the radiative decay are observed both theoretically and experimentally.     

Behaviour of the 3p 511s’ [1/2]1 level of Argon I in a strong magnetic field

The properties of the autoionizing 3p ⁵11s’ [1/2]₁ Argon I level have been experimentally studied in the presence of a strong magnetic field (2.14-2.22 T). Quasi- Landau type oscillations have been observed in the width and position of the 3p ⁵-4p[1/2]₀ -3p ⁵11s’ [1/2]₁ exciting transition. The results provide an additional support for earlier observations of Lemoigne et al. [7]. No evidence for similar oscillations in magnetic field dependence of the Fano q parameter was observed.     

The Stark broadening of the He I 667.8 nm line

The half-width and asymmetry parameter of the He I 2¹ P–3¹ D λ=667.8 nm line have been measured in a laser produced He plasma for electron densitiesN _e between 7×10²² m^?3 and 1.7×10²³ m^?3. Results, obtained for the half-width, agree very well with the theoretical results forN _e ?10²³ m^?3 and the agreement becomes somewhat worse forN _e >10²³ m^?3. We have measured the red asymmetry which is considerable lower than the theoretical one. We have also established good agreement with theoreticalj _A,R (x) profile for this line.     

Beam-foil study of doubly and singly excited states in BIII

Doubly excited⁴ L and² L systems of BIII have been studied by beam-foil spectroscopy. Spectra of boron (λ: 28.7–50.1 nm) have been recorded at Bochum with a resolution superior to that obtained previously. Eight new lines are classified in the⁴ L system and one in the² L system. The wavelength accuracy of most of the transitions already reported in the core-excited systems of BIII is greatly improved. On the basis of these results, six and two unidentified lines appearing in spectra recorded at Liège (λ: 50–200 nm) are classified in the⁴ L and² L core-excited systems, respectively. Where comparison with theoretical predictions by Chung et al. [1] is possible, excellent agreement is found for all the assigned lines. The first energy values are reported for seven terms in the doubly excited quartet system. Moreover, seven new lines are identified in the singly excited system of BIII.     

Solubility product and related thermodynamic quantities of silver and barium chromates in formamide

The values of ΔG⁰, ΔH⁰, and ΔS⁰ for the dissolution process, M₂(CrO₄)_x + solvent — 2 M^x+ (solvated) + xCrO₄^2? (solvated), where M is Ag or Ba, and x is 1 or 2, have been determined in formamide from solubility studies. The negative value of ΔS⁰ indicates that there is more order in the dissolved state than in the undissolved state.