Cryo-electron microscopy of freestanding molecules (single-particles) plays a pivotal role in the difficult and pressing challenge of determining the structures of large macromolecular complexes. Molecular volumes are generated by aligning large sets of randomly oriented two-dimensional (2D) projection images in three dimensions (3D) before reconstruction is performed using tomographic techniques. The increasing popularity of the single-particle method is highly correlated with technical advances in instrumentation and computation. This thesis introduces new computational methods for 3D structure determination from electron microscopic projection images of single molecules. The algorithms have been developed to fill a gap in the single particle methodology – the lack of methods for ab initio 3D reconstruction of asymmetrical or low-symmetry molecules co-existing in different functional states. The proposed approach does not rely on a priori information about the structure or the character of the sample heterogeneity, which minimizes template dependence and makes the methods applicable to a wide range of single molecules. The presented algorithms constitute the basis of a new open source software package - SIMPLE (Single-particle IMage Processing Linux Engine). SIMPLE is an efficient and easy-to-use image processing system for semi-automated ab initio 3D reconstruction from challenging single-particle data sets (asymmetrical particles, significant degree of heterogeneity). / QC 20101214

No description available.

<p>Molecular electronics is an active research area for the future information technology. The fabrication of basic electronic elements with molecules as the core-operators has been made experimentally in the laboratory in recent years. However, the underlying electron or charge transport mechanisms for most devices are still under debate, Theoretical modelling based on the first-principles methods are expected to play an important role in this field.</p><p>A generalized quantum chemical approach based on Green's function scattering theory has been developed and applied to two- and three-terminal molecular devices. It allows to study both elastic and inelastic electron scattering at hybrid density functional theory levels. It can treat molecular devices where the metal electrodes and the molecule are either chemically or physically bonded on equal footing. As one of the applications, we have studied the length dependence of electron transport in gold-oligophenylene-gold junctions. We have shown that the experimental results for molecular junctions of oligophenylene with di erent lengths can be well reproduced by hybrid density functional theory calculations. It is also found that the current-voltage characteristics of the junctions depend strongly on the metal-molecule bonding distances. With the help of the calculations, the possible gold-molecule bonding distances in the experimental devices are identi ed.</p><p>The central focus of this thesis is to study the three-terminal molecular devices, namely the eld e ect transistor (FET). An extension of our quantum chemical approach to FET devices has been made and successfully applied to different FET devices constructed with polymer, small and middle sized conjugated molecules. The experimentally observed conductance oscillation in polymer FET and three orders of magnitude enhancement of the current in electrochemical gated molecular FET have been verified by the calculations. The electron transport mechanisms of these devices are revealed.</p>

<p>The development of computational models for predicting drug solubility has increased drastically during the last decades. Nevertheless these models still have diffculties to estimate the aqueous solubility as accurate as desired. In this thesis di erent aspects that are known to have a large impact on the aqueous solubility of a molecule have been studied in detail using various theoretical methods with intension to provide microscopic view on drug solubility. The rst aspect studied is the hydrogen bond energies. Eight drug molecules have been calculated using density functional theory and the validity of additive model that has often been used in solubility models is examined. The impact of hydrogen bonds in Infrared and Raman spectra of three commonly used drug molecules has also been demonstrated. The calculated spectra are found to be in good agreement with the experimental data. Another aspect that is important in solubility models is the volume that a molecule occupies when it is dissolved in water. The volume term and its impact on the solvation energy has therefore also been calculated using three di erent methods. It was shown that the calculated volume di ered signi cantly dependent on which method that had been used, especially for larger molecules.</p><p>Most of the solubility models assume the solute molecule to be in the bulk of the solvent. The molecular behavior at the water/gas interface has been investigated to see how it di ers from bulk. It was seen that the concentration close to the interface was almost three times higher than in the bulk. The increase in concentration close to the surface depends on the larger gap between the interface energy and the gas phase energy than between the bulk energy and the gas phase energy.</p>

<p>The present thesis concerns theoretical studies of molecular interactions investigated by infrared and X-ray spectroscopic techniques, with emphasis on using these two techniques combined in pump-probe experiments. Four main types of studies are addressed: the use of near-edge X-ray absorption fine structure spectra (NEXAFS) to manifest through-bond and through-space interactions; the role of hydrogen bonding in the formation of X-ray photoelectron spectra as evidenced by simulations of the water dimer; the development of theory, with sample applications, for infrared X-ray pump-probe spectroscopy; and molecular dynamics simulations of light-induced fragmentation of water clusters.</p><p>Ab initio calculations indicate that NEXAFS spectra give direct information about the through-bond and through-space interactions between vacant non-conjugated π* orbitals. It is found out that the X-ray photoelectron spectrum of the water dimer differs dramatically from the monomer spectrum in that two bands are observed, separated by the chemically shifted ionization potentials of the donor and the acceptor. The hydrogen bond is responsible for the anomalously strong broadening of these two bands. The studies show that X-ray core electron ionization of the water dimer driven by an infrared field is a proper technique to prove the proton transfered state contrary to conventional X-ray photoelectron spectroscopy. </p><p>The physical aspects of the proposed new X-ray spectroscopic method - phase sensitive Infrared - X-Ray Pump-Probe Spectroscopy - are examined in detail using the wave packet technique in three applications; the NO molecule and the dynamics of proton transfer in core ionized water dimer and glyoxalmonoxime. It is found out that the phase of the infrared pump field strongly influences the trajectory of the nuclear wave packet on the ground state potential, which results in a phase dependence of the X-ray pump-probe spectra. A proper choice of the delay time of the X-ray pulse allows the direct observation of the X-ray transition in the proton transfered well of the core excited potential. It is found out that the glyoxalmonoxime molecule possesses an important feature; proton transfer accompanied by core hole hopping. Special attention is paid to the quantum control of the populations of vibrational level which is of crucial importance to shape the wave packet of desirable size.</p><p>The wave packet technique becomes computationally very expensive when the number of nuclear degrees of freedom is large. Molecular dynamics is used instead in studies of light-induced nuclear kinetics in the water hexamer cluster. We predict a novel mechanism of the mechanical action of light on atoms and molecules. This mechanism is based on the rectification of the Lorentz force, which gives a unique opportunity of direct site selective mechanical action of light on atoms and molecules inside large systems like clusters or biomolecules.</p>

<p>The work presented in the thesis concerned theoretical study of light-matter nonlinear interactions. Two important aspects of such interactions have been examined, namely the nonlinear optical properties of a series of organic charge transfer molecules in solutions induced by the laser light and the propagation of the ultrafast high power laser through the nonlinear molecular medium.</p><p>Special attention has been paid to understand the solvent effects on the two-photon absorption of a symmetrical diamino substituted distyrylbenzene chromophore, for which time-dependent density functional theory in combination with polarizable continuum model (PCM) have been employed. The dielectric medium alone has a rather small effect both on the bond length alternation and on the one-photon absorption spectrum, but noticeable effects on the two-photon absorption cross section. Both one- and two-photon absorptions are found to be extremely sensitive to the planarity of the molecule. Our calculations indicate that the experimentally observed anomalous solvent effect on the two-photon absorption of dialkylamino substituted distyrylbenzene chromophores can not be attributed to the intrinsic properties of a single molecule and its interaction with solvents. With the same theoretical approaches, two-photon absorption properties of interacting polar chromophores have been investigated to examine the validity of the widely used exciton model. Our first principles calculations have shown that the exciton model offers a conceptually simple interpretation for experimental observations, but is lack of predictability.</p><p>The second part of the thesis is to investigate the propagation of ultrashort laser pulse through a one-dimensional asymmetric organic molecular medium by solving full Maxwell-Bloch equations using predictor-corrector finite-difference time-domain method. It focuses on the supercontinuum generation of spectra and the formation of attosecond pulses. It is shown that the supercontinuum generation is strongly modulated by both area and width of the pulse, which results from the interference between the splitting pulses in time-domain and is the implication of the time-energy uncertainty relation. The presence of permanent dipole moment in molecular medium has noticeable effects on the supercontinuum generation. Our calculations show that a well-shaped 132 attosecond pulse can be generated from a two femtosecond incoming pulse under certain conditions. Influences of carrier-envelope phase and time-dependent ionization on the spectral and temporal evolutions of the ultrashort pulses have also been discussed.</p>

<p><i>In this thesis, quantum chemical methods are used to investigate enzymatic reaction mechanisms. The Density functional theory, in particular the hybrid B3LYP functional, is used to model two enzymes belonging to the tautomerase superfamily; 4-Oxalocrotonate Tautomerase (4-OT) and cis-Chloroacrylic Acid Dehalogenase (cis-CAAD). The methodology is presented and new mechanistic insights for the two enzymes are discussed.</i></p><p><i>For 4-OT, two different models are built and the potential energy curves are computed. This allows the methodology to be evaluated. The results give new insight into the energetics of the 4-OT reaction, indicating that the charge-separated intermediate is quite close in energy to the reactant species. The models also make it possible to perform in silico mutations to investigate the role of active site groups. Excellent agreement is found between the calculations and site-directed mutagenesis experiments, further substantiating the validity of the models.</i></p><p><i>For cis-CAAD, the uncatalyzed reaction is first considered and excellent agreement is found between the calculated barrier and the measured rate constant. The enzymatic reaction is then studied with a quite large active site model and a reaction mechanism is proposed.</i></p>

<p>The recombinant product is secreted to the periplasm in many processes where<em> E. coli</em> is used as host. One drawback with secretion is the undesired leakage of the periplasmic products to the medium.</p><p>The aim of this work was to find strategies to influence the periplasmic retention of recombinant products. We have focused on the role of the specific growth rate, a parameter that is usually controlled in industrial bioprocesses. The hypothesis was that the stability of the outer membrane in <em>E. coli </em>is gained from a certain combination of specific phospholipids and fatty acids on one side and the amount and specificity of the outer membrane proteins on the other side, and that the specific growth rate influences this structure and therefore can be used to control the periplasmic retention.</p><p>We found that is possible to control the periplasmic retention by the growth rate. The leakage of the product increased as the growth rate increased. It was however also found that a higher growth rate resulted in increased productivity. This resulted in equal amounts of product inside the cells regardless of growth rate.</p><p>We also showed that the growth rate influenced the outer membrane composition with respect to OmpF and LamB while OmpA was largely unaffected. The total amount of outer membrane proteins decreased as the growth rate increased. There were further reductions in outer membrane protein accumulation when the recombinant product was secreted to the periplasm. The lowered amount of outer membrane proteins may have contributed to the reduced ability for the cell to retain the product in the periplasm.</p><p>The traditional way to control the growth rate is through a feed of substrate in a fed-batch process. In this work we used strains with a set of mutations in the phosphotransferase system (PTS) with a reduced uptake rate of glucose to investigate if these strains could be used for growth rate control in batch cultivations without the use of fed-batch control equipment. The hypothesis was that the lowering of the growth rate on cell level would result in the establishment of fed-batch similar conditions.</p><p>This study showed that it is possible to control the growth rate in batch cultivations by using mutant strains with a decreased level of substrate uptake rate. The mutants also produced equivalent amounts of acetic acid as the wild type did in fed-batch cultivation with the same growth rate. The oxygen consumption rates were also comparable. A higher cell density was reached with one of the mutants than with the wild type in batch cultivations. It is possible to control the growth rate by the use of the mutants in small-scale batch cultivations without fed-batch control equipment.</p>

Density functional theory (DFT) has become the leading method in calculating theelectronic structures and properties from first principles. In practical applicationsof DFT in the frame work of Kohn-Sham (KS) method, an approximate exchange-correlation functional has to be chosen. Hence, the success of a DFT calculationcritically depends on the quality of the exchange-correlation functional.This thesis focuses on the development and validation of the so-called dou-bly hybrid density functionals (DHDFs). DHDFs present a new generation offunctionals, which not only have a non-local orbital-dependent component in theexchange part, but also incorporate the information of unoccupied orbitals in thecorrelation part. I will first give an overview of modern DFT in the introductorychapters, emphasizing the theoretical bases of a newly developed DHDF, XYG3.I will then present further examination of XYG3 and new development on top ofXYG3, leading to XYG3o and XYG3s. Attempts have also been made to extractband structure information of a periodic system from cluster model calculations. / QC 20110607

Microfluidic chips have become a powerful tool in research where biological cells are processed and/or analyzed. One method for contactless cell manipulation in microfluidic chips that has gained an increasing amount of attention the last decade is ultrasonic standing wave (USW) technology. This Thesis explores the biocompatibility of USW technology applied to microfluidic chips, and presents a novel USW-based method for serial processing and accurate characterization of living cells. The biocompatibility has been investigated by measuring the proliferation rate of cells after they had been trapped and aggregated inside a chip by ultrasound. No negative influence was observed after continuous exposure to 0.85 MPa pressure amplitudes for up to 75 min. Furthermore, the heat generation in the fluid channel caused by the ultrasound has been measured and used in a regulation scheme where the temperature can be controlled around any relevant temperature (e.g. 37‰) with ±0.1‰ accuracy for more than 12 hours. The proliferation rate and temperature investigations suggest that USW technology applied to microfluidic chips is a biocompatiblemethod useful for long-term handling of living cells. We have introduced a new concept of contactless ultrasonic ”caging” of single cells or small aggregates of cells. These cages are channel segments in the microfluidic chips that are geometrically designed to resonate at one or several actuation frequencies. The actuation is performed remotely by up to five external frequency specific wedge transducers, where each transducer produces a localized and spatially confined standing wave with a specific orientation of its corresponding radiation force field. By multi-frequency actuation, sophisticated and flexible force fields are realized by both overlapping and separated single fields. The Thesis describes two different cages: A sub-mm ”micro-cage” for tree-dimensional manipulationof single cells, and a 5-mm ”mini-cage” for selective retention of small cell aggregates (up to approx. 10^3 cells) from a continuously feeding sample flow. Finally,our microfluidic chips were also designed to be compatible with high-resolution optical microscopy. We have demonstrated sub-μm-resolution confocal fluorescence and trans-illumination microscopy imaging of ultrasonically caged living cells. / QC 20100811