Molecular mechanisms of crystallization and defect formation

Using the atomic force microscope (AFM) in situ during the crystallization of the protein apoferritin, we show that for this system the kink density along the steps is an equilibrium property that, multiplied by the frequency of molecular attachment, fully determines the propagation of growth steps. The intermolecular bond energy is 3.2k(B)T. Point defects are nonequilibrial and are caused by incorporation of impurity molecules, and replicate in subsequent layers due to the strain they cause. Using single-molecule manipulation with the AFM tip, the defects can be healed to restore the regular lattice.     

Evidence for non-DLVO hydration interactions in solutions of the protein apoferritin

We have studied molecular interactions in solutions of the protein apoferritin by static and dynamic light scattering. When plotted against the electrolyte concentration, the second osmotic virial coefficient exhibits a minimum. The ascending branch of this dependence is a manifestation of a surprisingly strong repulsion between the molecules at electrolyte concentrations about and above 0.2M, where electrostatic interactions are suppressed. We argue that the repulsion is due to the water structuring, enhanced by the accumulation of hydrophilic counterions around the apoferritin molecules, giving rise to so-called hydration forces.     

Attenuation of second sound in superfluid 3He-A1

The attenuation of second sound (spin-entropy) wave in the superfluid A1 phase has been measured in magnetic fields up to 11 T and to sufficiently high frequency to observe the bulk attenuation proportional to the square of frequency. The measured attenuation coefficient is compared with the existing theories of hydrodynamics and dissipative coefficients. The resulting "excess" attenuation is discussed in terms of the temperature dependent spin diffusion coefficient in the superfluid.     

Self-assembly of apoferritin molecules into crystals: thermodynamics and kinetics of molecular level processes

Aside from the biological, biomedical and materials importance of ferritin and apoferritin, the self-assembly of these molecules into crystals is of interest because it is a suitable model for protein crystallization and aggregation that impact many fundamental and applied fields. We use the atomic force microscope in-situ, during the crystallization of apoferritin to visualize and quantify at the molecular level the processes responsible for crystal growth. We image the configuration of the incorporation sites, “kinks”, on the surface of a growing crystal. We show that the kinks are due to thermal fluctuations of the molecules at the crystal-solution interface. This allows evaluation of the free energy of the intermolecular bond φ = 3.0 k_BT = 7.3 kJ/mol. The crystallization free energy, extracted from the protein solubility, is − 42 kJ/mol. Thermodynamics analyses based on these two values suggest that the main component in the crystallization driving force is the entropy gain of the waters bound to the protein molecules in solution and released upon crystallization. Furthermore, we determine the characteristic frequency of attachment of individual molecules into the kinks at one set of conditions. We show that the macroscopic step growth velocity, scaled by the molecular size, equals the product of independently determined kink density and attachment frequency, i.e., these are the molecular-level parameters for crystallization. Finally, we show that although new crystal layers are generated by intrinsically stochastic surface nucleation, for crystals > 200 μm layer generation predominantly occurs at the crystal edges. Numerical simulations of the concentration fields in the solution allow us to assign this localization to higher interfacial concentration at the edges. As the steps propagate to cover the crystal face, step density waves, or bunches, develop and may lead to non-uniformity and lower quality and utility of the growing crystal.     

Photoionization of Al IV (neon-type) from its ground 2s 22p 6 1 S e and excited 2s 22p 53p 3,1 P 0 3 S e states using the R-matrix method

The R-matrix method is used to calculate the total photoionization cross-sections from the ground 2s ²2p ^{6 1} S ^e and the first three excited 2s ²2p ⁵3p ^3,1 S ^e states of Al IV, for photon energies ranging from the first ionization threshold to just above the second threshold of the residual ion Al V. The two lowest LS terms of Al V − 2s ²2p ^{5 2} P ⁰, 2s2p ^{6 2} S ^e, 2s2p ^{6 2} S ^e, represented by sophisticated configuration interaction wavefunctions, are included in the R-matrix calculation. The resulting cross-sections are affected by Rydberg series of resonances converging to the 2s2p ^{6 2} S ^e excited threshold.     

Metal-sensitive and thermostable trypsin from the crevalle jack (<Emphasis Type="Italic">Caranx hippos</Emphasis>) pyloric caeca: purification and characterization

Background

Over the past decades, the economic development and world population growth has led to increased for food demand. Increasing the fish production is considered one of the alternatives to meet the increased food demand, but the processing of fish leads to by-products such as skin, bones and viscera, a source of environmental contamination. Fish viscera have been reported as an important source of digestive proteases with interesting characteristics for biotechnological processes. Thus, the aim of this study was to purify and to characterize a trypsin from the processing by-products of crevalle jack (Caranx hippos) fish.

Results

A 27.5 kDa trypsin with N-terminal amino acid sequence IVGGFECTPHVFAYQ was easily purified from the pyloric caeca of the crevalle jack. Its physicochemical and kinetic properties were evaluated using N-α-benzoyl-_DL-arginine-p-nitroanilide (BApNA) as substrate. In addition, the effects of various metal ions and specific protease inhibitors on trypsin activity were determined. Optimum pH and temperature were 8.0 and 50°C, respectively. After incubation at 50°C for 30 min the enzyme lost only 20% of its activity. K _m , k_cat, and k _cat /K _m values using BApNA as substrate were 0.689 mM, 6.9 s^-1, and 10 s^-1 mM^-1, respectively. High inhibition of trypsin activity was observed after incubation with Cd²⁺, Al³⁺, Zn²⁺, Cu²⁺, Pb²⁺, and Hg²⁺ at 1 mM, revealing high sensitivity of the enzyme to metal ions.

Conclusions

Extraction of a thermostable trypsin from by-products of the fishery industry confirms the potential of these materials as an alternative source of these biomolecules. Furthermore, the results suggest that this trypsin-like enzyme presents interesting biotechnological properties for industrial applications.

     

Deformation phase transition in vanadium under high pressure

Within the framework of the Landau theory of phase transitions and the density functional method, it is shown that the structural transformation from a body-centered cubic phase to a rhombohedral phase revealed in vanadium at 69 GPa is a first-order deformation phase transition close to a second-order phase transition.     

Nucleation of ordered solid phases of proteins via a disordered high-density state: phenomenological approach

Nucleation of ordered solid phases of proteins triggers numerous phenomena in laboratory, industry, and in healthy and sick organisms. Recent simulations and experiments with protein crystals suggest that the formation of an ordered crystalline nucleus is preceded by a disordered high-density cluster, akin to a droplet of high-density liquid that has been observed with some proteins; this mechanism allowed a qualitative explanation of recorded complex nucleation kinetics curves. Here, we present a simple phenomenological theory that takes into account intermediate high-density metastable states in the nucleation process. Nucleation rate data at varying temperature and protein concentration are reproduced with high fidelity using literature values of the thermodynamic and kinetic parameters of the system. Our calculations show that the growth rate of the near-critical and supercritical ordered clusters within the dense intermediate is a major factor for the overall nucleation rate. This highlights the role of viscosity within the dense intermediate for the formation of the ordered nucleus. The model provides an understanding of the action of additives that delay or accelerate nucleation and presents a framework within which the nucleation of other ordered protein solid phases, e.g., the sickle cell hemoglobin polymers, can be analyzed.