Acetate uptake by the unicellular cyanobacteria Synechococcus and Aphanocapsa.
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Biomedical subjects
Publications and source records attributed to J Gibson.
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The phenomenon of antigen-induced aggregation of human buffy coat leucocytes correlates well with skin testing using purified protein derivative of tuberculin as the antigen and in this system appears to be a simple, sensitive test of cell-mediated immunity. The same phenomenon may, however, be used as an indicator of humoral immunity. In addition antigen-induced buffy coat leucocyte aggregation is suppressed by serum from patients with Hodgkin's disease and sarcoidosis. The possible uses and also the limitations of this test are discussed.
An evaluation of methods for identification of Enterobacteriaceae was made employing the new commercial Micro-Media Enteric System (MMES) with that of the Analytab Products Incorporated (API) and the Conventional tube media schema as suggested by the Center for Disease Control (CDS). The MMES system employed 20 biochemical tests, the API 21, and the CDC procedure 25. Sixteen of these were identical biochemical tests. Two hundred clinical isolates of Enterobacteriaceae were tested employing procedures recommended by the manufacturers of MMES and API, and methods suggested by CDC. Among the sixteen identical biochemical tests the agreement was 98.0% (Conventional), 98.2% (API), and 97.98% (MMES). Bacteria misidentified by the API system totaled 5 (2.5%), 12 (6%) for the Conventional, and 13 (6.5%) for the MMES. Five of the bacteria misidentified with the MMES procedure were due to false positive citrate tests. This problem was subsequently eliminated. The results of this study indicated that the new MMES method for identification of Enterobacteriaceae compared favorably with both the API and Conventional procedures. However, significant advantages of the MMES method were evident in initial purchase price, utilization of technology time, and less tedium performing the test.
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Anacystis nidulans (Synechococcus) had a minimal doubling time of 5 hrs at 30 degrees C at saturating light intensity and carbon dioxide concentration. Half maximal growth rates in saturating CO2 occured at a light intensity of 0.54 mW per cm2, and there was an apparent threshold intensity of 0.13 mW per cm2 below which no growth occurred. Growth rate in saturating light was dependent on the concentration of CO2+H2CO3 in the medium, rather than on total dissolved CO2; half maximal rates were estimated at 0.1 mM CO2+H2CO3. Under saturating conditions of light and CO2, 14CO2 was fixed primarily into 3-PGA, and subsequently moved into sugar phosphates and amino acids. Incorporation into aspartate was relatively slow. CO2 fixation was strictly light-dependent. The changes in adenylate and pyridine nucleotide pools were followed in light/dark and dark/light transitions. Whereas adenylates relaxed slowly over 15-20 min to the concentrations characteristic of illuminated cells following the abrupt changes induced by darkening, the sharp drop in intracellular NADPH showed little dark recovery although rapid restoration occurred on reillumination. Other pyridine nucleotides showed no changes during these transitions. The nucleotide specificity and Km of partially purfied GAP dehydrogenase suggest a role for this enzyme in the regulation of CO2 fixation.
Anacystis nidulans (Synechococcus) was maintained in a medium of low phosphate concentration (0.1 mM) and grew with a normal doubling time of 5 hrs at 30 degrees C. Such cultures had a normal pigment composition and alkaline phosphatase was detectable at low specific activities only. The onset of phosphate-limited growth occurred when the phosphate concentration in the medium fell to a value below 4 muM (the limit of accurate determination by the assay method used ) and resulted in increases in alkaline phosphatase activity, reaching a final 10 to 15 fold increase in specific activity after a period of several hours. Marked changes in the overall pigment composition occurred in this period of growth restriction. The addition of phosphate to such cultures resulted in a halt in synthesis of the enzyme and the restoration of normal pigmentation before growth resumed at the normal rate. Several oraganic phosphate esters could replace inorganic phosphate for growth and were also hydrolyzed by the partially purified enzyme, but growth rates were characteristically lower and the specific activity only 3 to 4 fold higher than in cultures grown in phosphate excess. Studies with the partially purified enzyme suggested that it differed in some of its properties from other alkaline phosphatases described in the literature.
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The uptake of C4 dicarboxylates by cells from exponential cultures of Rhodopseudomonas spheroides followed saturation kinetics at concentrations below 100 muM with Km values for succinate, malate, and fumarate of 2.7, 2.3, and 0.8, respectively. Corresponding Vmax values of 50, 52, and 67.5 nmol/min per mg of protein at 20 C were obtained. Each of these compounds interfered competitively with uptake of the others, and a common transport system appears to be involved. Fructose-grown cells took up C4 dicarboxylates only at very low rates, and pyruvate-grown cells took up C4 dicarboxylates at one-third the rates found with succinate-grown cultures. Malonate and maleate inhibited uptake less severely, and aspartate and alpha-ketoglutarate had no effect at 100-fold excess. Divalent metals stimulated uptake. Light or respiration was required for uptake, and entered materials were rapidly converted to other metabolities, notably amino acids. Pyruvate entry appeared to be mediated by several systems, of which only one could be resolved kinetically. This system had a Km of 13 muM and Vmax of 5.6 nmol/min per mg of protein at 20 C. A number of related mono- and dicarboxylates interfered with pyruvate uptake. The pyruvate uptake system was distinguishable from the C4 dicarboxylate system by the absence of divalent cation stimulation and by substrate and inhibitor specificity.
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Adenine nucleotide pools and their energy charge were measured during balanced and unbalanced growth of photoheterotrophic Chromatium cultures. The methods used involved rapid sampling, accurate to within 1 s, from isotopically labeled cultures followed by chromatographic separation of individual nucleotides. During balanced growth, both energy charge and adenosine triphosphate (ATP) concentrations, whether expressed as a function of cell protein or intracellular water, were slightly higher in limiting light intensities than in cultures growing at their maximal rate in bright light. The ATP found corresponded to 4.67 +/- 0.08 nmol/mg of protein or 1.34 +/- 0.57 mM for low-light cells and to 4.41 +/- 0.58 mmol/mg of protein or 0.85 +/- 0.12 mM for high-light cells. Corresponding energy charges were 0.85 +/- 0.02 and 0.81 +/- 0.02. Illumination shifts caused differential synthesis of photosynthetic pigments lasting 2 to 3 h without corresponding perturbation of adenine nucleotide levels. Cultures in intermittent illumination were severely affected by some cycle durations; they had abnormal morphology and very high bacteriochlorophyll-to-protein ratios. In such cultures, energy charge and nucleotide concentrations were within normal limits and relaxed to the dark steady state during the dark periods. Arsenate at AsO(4) (3-) to PO(4) (3-) ratios of 10:1 in the medium retarded growth, but no abnormality of charge or quantity of phosphate-containing nucleotides was found. These experiments therefore suggest that, within experimental error, neither the size nor the charge of the adenylate pools governs growth rate in Chromatium. Moreover, these parameters do not appear to be concerned in regulating the synthesis of photosynthetic apparatus in this organism.
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