Ionization behavior of the histidine residue in the catalytic triad of serine proteases

α-Lytic protease is a homologue of the mammalian serine proteases such as trypsin, chymotrypsin, and elastase, and its single histidine residue belongs to the Asp-His-Ser catalytic triad. This single histidine residue has been selectively enriched in the C-2 carbon with 13C. Magnetic resonance studies of the chemical shift and coupling constant (1Jch) behavior of this nucleus as a function of pH suggest that the imidazole ring is neutral above pH 5 and therefore that the group which is known to ionize with pKa near 6.7 must be the aspartic acid residue. Implications of these new pKa assignments for the catalytic mechanism of serine proteases are discussed and include the absence of any need to separate charge during catalysis. The histidine residue plays two roles. (a) It insulates the aspartic acid from an aqueous environment and accordingly raises its pKa. (b) It serves as a bidentate base to accept a proton from the serine at one of its nitrogens and concertedly transfer a proton from its other nitrogen to the buried carboxylate anion during formation of the tetrahedral intermediate

In-beam internal conversion electron spectroscopy with the SPICE detector

The SPectrometer for Internal Conversion Electrons (SPICE) has been commissioned for use in conjunction with the TIGRESS $\gamma$-ray spectrometer at TRIUMF's ISAC-II facility. SPICE features a permanent rare-earth magnetic lens to collect and direct internal conversion electrons emitted from nuclear reactions to a thick, highly segmented, lithium-drifted silicon detector. This arrangement, combined with TIGRESS, enables in-beam $\gamma$-ray and internal conversion electron spectroscopy to be performed with stable and radioactive ion beams. Technical aspects of the device, capabilities, and initial performance are presented

Structural studies of the recognition of bacterial lipopolysaccharides by human surfactant protein-D

SP-D is a large hydrophilic protein, consisting of four collagenous trimers, which is secreted by alveolar type II and non-ciliated bronchiolar epithelial cells and participates in calcium-dependant agglutination of inhaled microorganisms. Known high-resolution crystal structures of mono- and disaccharide bound recombinant human SP-D include a heptose disaccharide that mimics the inner core fragment of bacterial lipopolysaccharide. This thesis focuses on the structural characterisation of SP-D binding to lipopolysaccharides from Gram-negative bacteria. Intact LPS and several hydrolysed smooth and rough mutants were soaked into native crystals of a recombinant head and neck fragment of SP-D. Those soaked with hydrolysed Escherichia coli J5 and Salmonella minnesota R7 LPS showed electron density corresponding to ligand in the ligand-binding site. All crystals processed conformed to spacegroup P21, all with unit cells close to a= 55, b= 108, c= 55 Å, = 91°. The maximum resolution diffraction observed was 1.63 Å. The R7-soaked structure was refined at 1.77 Å, with a final R-factor of 18.71% and R-free of 22.05%. The structure reveals a well-defined trisaccharide unit of two heptose saccharides and a single Kdo residue in protein chains B and C. The Kdo is present as a five-membered, anhydro residue known to form during mild acid hydrolysis. The third, outermost heptose of the R7 inner core is not visible in the electron density. The refined structure demonstrates binding of LPS via the O6 and O7 hydroxyl groups of the glycerol sidechain of HepI, the innermost heptose, demonstrating for the first time structurally not only the binding of a physiologically relevant bacterially derived ligand but also the recognition of a non-terminal monosaccharides by SP-D. No direct interaction is observed between the second heptose and the protein, but two hydrogen bonds are seen between the anhydro-Kdo and the amino acids Arg343 and Asp325 which flank the SP-D ligand binding pocket

High-intensity interval training

High-intensity interval training (HIIT) is characterised by brief, intermittent bursts of near- or maximal-intensity exercise, interspersed by periods of active or passive recovery. The limited available evidence suggests that HIIT is an efficacious training method for young athletes. The effect of HIIT on cardiorespiratory fitness (CRF), endurance performance, explosive strength and sport-specific performance has been examined in a range of young athletic populations from various sports. Furthermore, promising preliminary findings suggest that HIIT may confer further benefits to a range of health outcome measures including fasting insulin, lipoproteins, systolic blood pressure and endothelial function; obese youth may benefit particularly from this type of training. Improved cardiorespiratory fitness has been observed consistently after HIIT in athletic and non-athletic populations. Larger studies, extended over longer periods, that include valid measures of exercise compliance, tolerance and enjoyment are required to further delineate the priority that could be afforded to this type of training

Modern NMR Pulse Sequences in Pharmaceutical R & D

NMR pulse sequences are surveyed for solution-state methods that serve as typical, robust techniques in pharmaceutical or chemical NMR laboratories. Attention is drawn to up-to-date methods capable of enhancing sensitivity, resolution, and information content. Sequences range from those used for pulse calibration and field homogeneity adjustment, through one- and two-dimensional homo- and heteronuclear methods for solution-phase work. Techniques used for editing, resolving, and simplifying data are highlighted and extensive use is made of sequence diagrams to present the basic structure of each pulse sequence in pictorial form. Where appropriate, descriptions of each sequence and some examples of data are provided and attention drawn to the advantages of using each technique

Role of Multichance Fission in the Description of Fission-Fragment Mass Distributions at High Energies

Fission-fragment mass distributions were measured for U237-240, Np239-242, and Pu241-244 populated in the excitation-energy range from 10 to 60 MeV by multinucleon transfer channels in the reaction O18+U238 at the Japan Atomic Energy Agency tandem facility. Among them, the data for U240 and Np240,241,242 were observed for the first time. It was found that the mass distributions for all the studied nuclides maintain a double-humped shape up to the highest measured energy in contrast to expectations of predominantly symmetric fission due to the washing out of nuclear shell effects. From a comparison with the dynamical calculation based on the fluctuation-dissipation model, this behavior of the mass distributions was unambiguously attributed to the effect of multichance fission