Resolution and reconstitution of glutamate decarboxylase from cerebellum

Brain glutamate decraboxylase (EC 4.1.1.15) catalyzes the biosynthesis of the postulated neurotransmitter-aminobutyric acid according to the following chemical equation:L-glutamate -aminobutyric acid+CO₂. Hydroxylamine treatment of the decarboxylase at low ionic strength followed by Sephadex gel filtration resolves apoenzyme from cofactor (>90%). Pyridoxal phosphate completely restores activity. Sodium borohydride inactivates the holoenzyme, but not the apoenzyme. This supports the notion that pyridoxal phosphate is bound to the holoenzyme as a Schiff base. Moreover, salicylaldehyde, a reagent which reacts with amino groups, substantially inactivates the apoenzyme but not the holoenzyme. Reconstitution of the bovine cerebellar holoenzyme from apoglutatamate decarboxylase and pyridoxal phosphate occurs in seconds to minutes, which is much faster than that of the decarboxylase isolated fromE. coli. Native holoenzyme, apoenzyme, and reconstituted holoenzyme have identical molecular weights as estimated by Sephadex gel filtration.A preliminary account of this work has been presented (1).     

Laser light-scattering investigations of the teleost swimbladder response to acoustic stimuli.

When a laser pencil is directed through the teleost swimbladder fringe patterns can be seen in the far-field that are (a) highly sensitive to the orientation and position of the swimbladder with respect to the incident pencil and (b) a representation of contributions from each membrane through which the light passes. The fringe pattern fluctuates in intensity, and to some extent in position, in response to driving forces that distort the swimbladder. The spectrum of these very small distortions can be measured by standard light scattering techniques. This method was used to study the response of in situ swimbladders to imposed acoustic fields and evidence for a sharp roll-off of the response at frequencies above 1,000 Hz was found. Models for these effects are discussed.     

Studies on rabbit lymphocytes in vitro : XX. Demonstration of cells reactive to more than one mitogen by mixed sequential stimulation

Rabbit peripheral blood lymphocyte (PBL) cultures stimulated by ConA and then blocked by the addition of competing sugar or antiserum after 6–15 h of ConA prestimulation respond to restimulation by PHA or PWM to a much greater extent than to continuous stimulation or delayed stimulation with PHA or PWM. This effect of mixed lectin sequential stimulation indicates that many of the same PBLs will respond to more than one mitogen, but that some cells require preactivation by one mitogen in order to respond fully to another mitogen. Thus, the number of PBLs which respond to PHA or PWM alone is much less than the number which respond following pretreatment with ConA when the pretreatment effect of ConA alone is blocked. Rabbit PBLs do not respond to LPS and preactivation by ConA does not prepare rabbit PBLs to respond to LPS.     

Some effects of light on the energetics of Daphnia pulex and implications for the significance of vertical migration

Energy budgets were computed from data obtained for Daphnia pulex cultured under nine light intensities, polarized light and four wavelength ranges. The percent assimilation of preadult animals is highest at intensities above 7 ft-c. Net efficiency of growth was highest (> 50%) and the net efficiency of respiration was lowest ( < 49%) at intensities less than 28 ft-c. The percent assimilation of adult animals was highest ( > 10%) at 110, 55 and 14 ft-c. Under the nine intensities the gross efficiencies of growth were less than 1 % and net efficiencies of growth varied from 3.9 to 7.3%. Gross efficiencies of respiration were highest above 7 ft-c. The net efficiency of respiration usually varied between 20 and 30% and the lowest was 9.8% at 1.7 ft-c. and the highest was 50.1% at 110 ft-c. Gross efficiency of reproduction varied from 2.6% at 3.5 ft-c to 12.6% at 14 ft-c and generally varied between 4 and 7.5%. Net efficiency of reproduction varied from 45.9% at 110 ft-c to 84.3% at 1.7 ft-c and usually varied from 62 to 75% at other light intensities. The ratio of energy of respiration to energy of growth and reproduction ranged from 12% to 1.7 ft-c to 105.3% at 110 ft-c. This ratio usually varied from 25 to 34% at 14 ft-c or less and exceeded 37% at intensities above 14 ft-c. The percent assimilation (3.5%), gross (2.0%), and net (56.3%) efficiencies of respiration of preadult animals raised under polarized light were higher than for those at a similar, nonpolarized, intensity. The net efficiency of growth (43.7%) was lower under polarized light. The percent assimilation, gross efficiencies of growth, reproduction and respiration, net efficiencies of growth and reproduction of adult animals under polarized light (6.6 ft-c) were lower than for those under 7 ft-c. For preadult animals assimilation efficiencies were lower in wavelength treatments than in white light or darkness. The gross efficiencies of growth and respiration were lowest under red wavelengths and the net efficiencies of growth were lowest and respiration highest under green wavelengths. For adult animals, the assimilation efficiencies were lower in the wavelength treatments than those obtained in other light treatments. While the gross efficiencies of growth, reproduction and respiration were generally lower, the net efficiencies of growth and reproduction were generally within the range of values for other light conditions. The net efficiencies of respiration, except for red wavelengths, were lower than those for other light conditions except at 1.7 ft-c. The ratio of energy of respiration to energy of growth and reproduction showed similar trends. The effects of wavelength are generally separable from the effects of light intensity.     

Incidence of Aflatoxin in California Almonds

In a survey of California almonds, aflatoxin was found in 14% of 74 samples of unsorted, in-shell almonds as received by the processor in 1972, but it occurred at very low levels (below 20 parts per billion [ppb]) in 90% of the contaminated samples. The overall proportion of individual nuts contaminated was especially low and is estimated with 95% probability to have been in the range of 1 nut/55,300 nuts to 1 nut/14,700 nuts. Aflatoxin contamination is not restricted to any particular section of the almond-growing region of California. Commercial sorting procedures are effective in removing most aflatoxin-contaminated nutmeats, since none of 26 samples of processed, whole nutmeats contained aflatoxin. In contrast, 13 of 27 samples of diced almonds were contaminated, but nine of these 13 samples contained less than 20 ppb. Only one of 25 samples of sliced nutmeats contained aflatoxin (4 ppb). Thus, aflatoxin incidence in almonds varies greatly with the category of finished product. The apparent high incidence in diced nutmeats is probably due mostly to the more uniform distribution of aflatoxin occurring in this product (because of its small particle size) than that occurring in the other products. Sample size requirements for monitoring aflatoxin in almonds are discussed.     

Distribution and effects of 2,4,5-trichlorophenoxyacetic acid in cells of Bacillus megaterium.

Cell death in a resting population of an asporogenous Bacillus megaterium was accelerated by ambient concentrations of 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) equal to or greater than 10 mug/ml or 5 mug/mg of cells (dry weight), but only after prolonged exposure. Conversely, populations of growing cells were not markedly influenced even at 100 mug/ml. Effects on cell respiration were not manifest until the ambient concentration reached 1,000 mug of 2,4,5-T/ml, or 500 mug/mg. Cells of B. megaterium did, however, accumulate 2,4,5-T passively to a level approximately twofold above the ambient concentration. Most of the accumulated compound was easily washed from the cells, but, of the firmly bound herbicide, about 0.5 mug/mg of cells (dry weight), nearly 60% by weight, was localized in the protoplast membrane. The foregoing results, obtained with a purified preparation of 2,4,5-T were also elicited by 2,4,5-T analytical standards. The extracted contaminants did not produce the results alone nor did they influence the results when present in combination with 2,4,5-T.     

Persistence and Biodegradation of Spilled Residual Fuel on an Estuarine Beach

The enrichment of hydrocarbon-degrading bacteria and the persistence of petroleum hydrocarbons on an estuarine beach after a spill of residual fuel oil on 11 April 1973 in Upper Narragansett Bay, R.I. was investigated. A rapid enrichment occurred during days 4 to 16 after the oil spill and a significant population of hydrocarbon-degrading bacteria was maintained in the beach sand for at least a year. The concentration of petroleum hydrocarbons in the mid-tide area declined rapidly during the bacterial enrichment period, remained fairly constant throughout the summer, and then declined to a low concentration after 1 year. An increased concentration of branched and cyclic aliphatic hydrocarbons in the low-tide sediment 128 days after the spill suggested a migration of hydrocarbons during the summer. Hydrocarbon biodegradation was apparent during the winter months at a rate of less than 1 mug of hydrocarbon per g of dry sediment per day.     

Lipid metabolism during bacterial growth, sporulation, and germination: an obligate nutritional requirement in Bacillus thuringiensis for compounds that stimulate fatty acid synthesis.

The regulation of fatty acid biosynthesis by compounds that are required for growth of Bacillus thuringiensis was investigated using an vivo assay developed to measure fatty acid synthesis in germinating spores. A minimal glucose-ammonium-salts medium does not support growth even though previous radiorespirometric studies have shown B. thuringiensis to possess intact tricarboxylic acid and Embden-Meyerhof-Parnas pathways. Abundant growth does occur, however, when this medium is supplemented with either glutamate, aspartate, citrate, thiosulfate, cystine, or ethylenediaminetetraacetic acid. Cells held under nongrowing conditions incorporate acetate into fatty acids; fatty acid synthesis is stimulated by the compounds that permit growth. These alternate nutritional requirements are not manifestations of a vitamin or trace metal deficiency and do not reflect a chelation phenomenon. These results indicate a direct correlation between the capacity of these compounds to promote growth and to stimulate formation of fatty acids.     

Soluble and membrane-bound aspartate-binding activities in Salmonella typhimurium.

The specificities of the soluble and membrane aspartate-binding activities were compared with each other and with the specificity of aspartate chemotaxis and were found to be distinct. The soluble aspartate-binding protein was purified to homogeneity and had a molecular weight of 30,000. The dissociation constant was 10(-6) M for aspartate, and the protein bound glutamate, cysteic acid, and 2-amino-3-phosphonopropionate. Aspartate transport was inhibited by cysteic acid.