Identification of neutral and anionic 8 alpha-substituted flavin semiquinones in flavoproteins by electron spin resonance spectroscopy

Circular dichroic spectra of metmyoglobin and apomyoglobin were measured in neutral and acidic solution. Addition of sodium dodecyl sulfate (NaDodSO₄) slightly reduces the helicity (based on the circular dichroic magnitude) of both proteins probably because of the loss of long-range interactions among helical segments. Lowering the pH of the protein-surfactant solution to 3 slightly enhances the helical conformation of myoglobin due to the protonation of acidic side groups and thereby the reduction of coulombic repulsion among negative charges. For BrCN-digested fragments the COOH-terminal peptide (22 residues) loses its helicity which can be restored by addition of NaDodSO₄. The middle fragment (76 residues) retains a considerable amount of helicity in water alone, which further increases in the presence of NaDodSO₄. The NH₂-terminal fragment (55 residues) also has some helical conformation in water, which is enhanced by the addition of NaDodSO₄. The circular dichroic spectrum of an equimolar mixture of the three peptides in NaDodSO₄ solution is the same as that calculated from the spectra of isolated peptides under similar conditions.     

Relation of phospholipase D activity to the decay of succinate dehydrogenase and of covalently bound flavin in yeast cells undergoing glucose repression

The disappearance of succinate dehydrogenase activity and of protein-bound histidyl flavin were studied in aerobic yeast cells incubated with high glucose concentrations. The decay of succinate dehydrogenase activity, covalently bound flavin, and of respiration is prevented by cycloheximide but not by chloramphenicol. During this decay there is a large increase in mitochondrial phospholipase D activity; the appearance of this enzyme is also prevented by cycloheximide. It seems possible, therefore, that the formation of phospholipase D may be important in triggering the disappearance of covalently bound flavin, succinate dehydrogenase, and of other mitochondrial enzymes during glucose repression of aerobic yeast cells.     

Human ankle joint stiffness over the full range of muscle activation levels

System identification techniques have been used to track changes in dynamic stiffness of the human ankle joint over a wide range of muscle contraction levels. Subjects lay supine on an experimental table with their left foot encased in a rigid, low-inertia cast which was fixed to an electro-hydraulic actuator operating as a position servo. Subjects generated tonic plantarflexor or dorsiflexor torques of different magnitudes ranging from rest to maximum voluntary contractions (MVC) during repeated presentations of a stochastic ankle angular position perturbation. Compliance impulse response functions (IRF) were determined from every 2.5 s perturbation sequence. The gain (G), natural frequency (omega n), and damping (zeta) parameters of the second-order model providing the best fit to each IRF were determined and used to compute the corresponding inertial (I), viscous (B) and elastic (K) stiffness parameters. The behaviour of these parameters with mean torque was found to follow two simple rules. First, the elastic parameter (K) increased in proportion to mean ankle torque as it was varied from rest to MVC; these changes were considerable involving increases of more than an order of magnitude. Second, the damping parameter (zeta) remained almost invariant over the entire range of contractions despite the dramatic changes in K.     

Partial IgA-deficiency with increased Th2-type cytokines in TGF-beta 1 knockout mice.

Though it has been shown that TGF-beta 1 directs B cells to switch to IgA in vitro, no studies have assessed TGF-beta 1 effects on mucosal vs systemic immunity in vivo. When the B cell functions of TGF-beta 1 gene-disrupted (TGF-beta 1-/-) mice were analyzed, significantly decreased IgA levels and increased IgG and IgM levels in serum and external secretions were observed. Further, analysis of Ab forming cells (AFC) isolated from both mucosal and systemic lymphoid tissue showed elevated IgM, IgG, and IgE, with decreased IgA AFC. A lack of IgA-committed B cells was seen in TGF-beta 1-/- mice, especially in the gastrointestinal (GI) tract. Splenic T cells triggered via the TCR expressed elevated Th2-type cytokines and, consistent with this observation, a 31-fold increase in serum IgE was seen in TGF-beta 1-/- mice. Thus, uncontrolled B cell responses, which include elevated IgE levels, a lack of antiinflammatory IgA, and an excess of complement-binding IgG and IgM Abs, will promote inflammation at mucosal surfaces in TGF-beta 1-/- mice and likely contribute to pulmonary and GI tract lesions, ultimately leading to the early death of these mice.     

Stage‐dependent physiological responses in a butterfly cause non‐additive effects on phenology

Studies illustrating climate‐induced shifts in phenology typically focus on the timing of a single lifecycle stage. In contrast, species’ responses to climate change are likely to be complex and constrained by interactions and tradeoffs across the lifecycle. We characterized the thermal sensitivity of egg, larval and pupal stages of a native Australian butterfly and then integrated these responses to predict sensitivity of emergence time, survival, and feeding performance on oviposition date and climate. Thermal physiology varied among lifecycle stages and between sexes, with the development rate of eggs, first instar larvae, and pupae being the most sensitive to temperature. As lifecycle stages have different thermal physiologies, the environment experienced by a given stage depends in a complex way on the experience of previous stages. Our simulations indicate that oviposition date strongly influences time spent in each lifecycle‐stage, as well as performance. Under a high emissions climate warming scenario (CSIRO Mk 3.5 climate model, high emissions SRES marker scenario A1F1, and a moderate rate of global warming), we predict development times to decrease by 38 days by 2070. Our analysis illustrates how differences in thermal physiology across the lifecycle may result in non‐additive effects on phenology which, in turn, may constrain species’ responses to global warming. These results highlight the need to view shifts in phenology in the context of an organism's entire lifecycle.     

Recovery of dynamic muscular endurance

Recovery of the rate of dynamic muscular endurance was measured in two groups of college-aged males. Subjects were required to perform elbow flexion (between the angles of 70 and 170 degrees) for as long as possible at the rate of 38 contractions/min while loaded with 1/6 of their maximum isometric strength (MVC). The task was terminated when the subject fell four contractions behind the required cadence or failed to complete two successive contractions. Subsequent to the task the subject was given a predetermined rest period after which a second fatigue bout to failure was performed. The rest intervals for Gp I (n = 22) were 5, 15, 45, 135, 405, and 1215 seconds, while the rest intervals for Gp II (n = 17) were 10, 30, 90, 270, 810, and 2550 s. Each subject completed six recovery intervals with the order of administration assigned at random. The percentage of recovery was calculated by dividing the exercise time of the first bout into the time of the second bout. These normalized data for the two groups were combined for analysis providing a 12 point recovery curve. The percentage of recovery ranged from 15.4% after 5 s to 91.8% after 2550 s. Analysis of the data revealed that the recovery pattern of dynamic muscular endurance progressed very rapidly initially, reached 50% at approximately 2 min and 15 s and was slightly less than 90% complete at 20 min. Exponential analysis of these data yielded a three-component curve.     

Methods for peptide identification by spectral comparison

Background  

Tandem mass spectrometry followed by database search is currently the predominant technology for peptide sequencing in shotgun proteomics experiments. Most methods compare experimentally observed spectra to the theoretical spectra predicted from the sequences in protein databases. There is a growing interest, however, in comparing unknown experimental spectra to a library of previously identified spectra. This approach has the advantage of taking into account instrument-dependent factors and peptide-specific differences in fragmentation probabilities. It is also computationally more efficient for high-throughput proteomics studies.     

Azabenzthiazole inhibitors of leukotriene A4 hydrolase

Previously, benzthiazole containing LTA₄H inhibitors were discovered that were potent (1–3), but were associated with the potential for a hERG liability. Utilizing medicinal chemistry first principles (e.g., introducing rigidity, lowering c Log D) a new benzthiazole series was designed, congeners of 1–3, which led to compounds 7a, 7c, 12a–d which exhibited LTA₄H IC₅₀ = 3–6 nM and hERG Dofetilide Binding IC₅₀ = 8.9–> >10 μM.