Pyridoxal phosphate-sensitized photoinactivation of glutamate decarboxylase from Clostridium perfringens

1. l-Glutamate decarboxylase (EC 4.1.1.15) from Clostridium perfringens was inactivated by exposure to visible light at pH6.2. 2. Inactivation does not occur at pH4.6 or in the absence of bound pyridoxal phosphate. 3. On prolonged photo-oxidation six histidine residues per molecule of enzyme were destroyed. 4. The loss of six cysteine residues per molecule occurred both in irradiated samples and in controls oxygenated in the dark. 5. This dark-oxidation of cysteine residues is apparently required before the photo-oxidation process. 6. The absorbance, fluorescence and circular-dichroism properties of the enzyme as well as its elution volume during Sephadex gel-filtration were unaffected by prolonged irradiation. 7. However, an apparently homogeneous product of photo-oxidation could be separated from the control enzyme by ion-exchange chromatography. 8. The K(m) for l-glutamate was unchanged in an irradiated sample retaining 22% of control activity. 9. These data and the catalytic role of imidazole residues at the active sites of amino acid decarboxylases are discussed.     

Genetic analysis of a population of Tribolium. V. Viability of the unsaturated fatty acid sensitive mutant.

The viability pattern of the unsaturated fatty acid sensitive mutant (cos/cos) of Tribolium castaneum was examined with reference to the normal genotype from egg hatching through adult emergence. The beetles were cultured on diets with 0, 3, or 5% corn oil in growth chambers maintained at 33 +/- 1 degree C and either 42 +/- 6% or 75 +/- 3% relative humidity. The pattern of the mutant was altered during the larval stage and was dependent on both the concentration of dietary fatty acid and the relative humidity of the culture chamber. Values of the viability component of fitness were assigned to the cos/cos, +/-/cos and +/-/+/- genotypes for multiple environments based on the number of eggs surviving to pupation. In large, random mating populations segregating at this locus, a stable genetic polymorphism is forecast. Predicted rates of change of allele frequency are dependent upon the culture conditions.     

Measurement of monomer-oligomer distributions via fluorescence moment image analysis

We present higher-order moment analysis of fluorescence intensity fluctuations from individual laser scanning microscopy images applied to study monomer-oligomer distributions. We demonstrate that the number densities and brightness ratios of a mixed population of monomers and oligomers can be determined by analyzing higher-order moments of the fluorescence intensity fluctuations from individual images for specific ranges of densities and particle brightness ratios. Computer simulations and experiments with fluorescent microspheres and cells were performed to illustrate the detection limits and accuracy of this statistical approach. The simulation results show that the concentration of the dimer or oligomer population should be less than or equal to the monomeric concentration for the method to provide accurate results, and that the upper density detection limit of the population of monomers is one order-of-magnitude higher than the concentration of the oligomers. We implemented this technique to resolve two populations of fluorescent microspheres with different brightness ratios and we also applied the moment-analysis method to examine the distribution of aggregation states of PDGF-beta receptors in human fibroblast cells. The method was able to resolve a tetrameric population of the PDGF-beta receptors relative to the background distribution of nonspecifically bound fluorophore.     

Natural selection and fitness entropy in a density-regulated population

The entropy H(p_o,p*) of a population with the initial allele frequency p_o given the equilibrium polymorphic frequency p* has been proposed as a measure of natural selection. In the present paper, we have extended this concept to include a particular aspect of density-dependent selection. We compared size trajectory of a population initially at genetic equilibrium, N(t), with the size trajectories of populations not initially at p*,N(t), but which do eventually converge to a common equilibrium allele frequency and equilibrium density, N*. The following experimentally testable hyopthesis was established. The total area defined by the difference between the trajectories of N(t) and N(t) as they converge to N* is directly proportional to the fitness entropy when population size is transformed using the density-dependent fitness value. Two properties of this relationship were noted. First, it is independent of the magnitude of natural selection and, secondly, it does not depend upon the initial population density as long as the equilibrium and nonequilibrium populations have the same initial numbers. This hypothesis was evaluated with experimental data on the flour beetle Tribolium castaneum.     

Experimental evidence suggesting that nitric oxide diffuses from tissue into blood but not from blood into tissue

The aim of this study was to evaluate in vivo whether nitric oxide (NO) is able to diffuse from blood into tissues and vice versa from tissues into blood. We used an in vivo model of intestinal ischemia (superior mesenteric artery occlusion) selectively increasing NO levels in intestinal tissue and an infusion of L-arginine selectively increasing NO levels in blood. In this model we followed formation of nitrosyl complexes of hemoglobin (Hb-NO) in blood and nitrosyl-diethyldithiocarbamate-iron complexes (DETC--Fe--NO) in ischemic intestine and normoxic tissues by means of electron paramagnetic resonance spectroscopy. NO trapping by DETC--Fe in the tissues resulted in a reduction of Hb--NO levels in blood accompanied by the formation of water-insoluble DETC--Fe-NO complexes in ischemic intestine and normoxic tissues both during ischemia and during reperfusion. Administration of L-arginine increased NO levels in blood but neither in ischemic intestine nor in normoxic tissue. Our data suggest that NO released in blood from endothelial cells does not diffuse into tissue. In contrast, NO formed in tissue diffuses into blood. The latter indicates that NO formed in tissues may exert its biological activities systematically.     

Explaining and predicting patterns in stochastic population systems

Lattice effects in ecological time-series are patterns that arise because of the inherent discreteness of animal numbers. In this paper, we suggest a systematic approach for predicting lattice effects. We also show that an explanation of all the patterns in a population time-series may require more than one deterministic model, especially when the dynamics are complex.