Biomedical subjects
Publications and source records attributed to C B Braun.
Pseudomonas sp. strain Kim has previously been reported to be the only known naturally occurring organism lacking spermidine. We now show that it synthesizes this polyamine. The apparent lack of intracellular levels of spermidine results from an efficient conversion of spermidine to putrescine and hydroxyputrescine.
We have previously shown that serum levels of C1q, unbound to C1r X C1s, are elevated in rheumatoid arthritis. We have also shown that RHP, a newly described serum protein which affects the C1q-anti C1q precipitin reaction, is also present at elevated levels in rheumatoid arthritis. We now show that RHP inhibits the hemolytic activity of C1q, disaggregates C1, and inhibits the ability of C1q bound to latex beads or to aggregated IgG to enhance the oxidative metabolism of neutrophils.
The radial immunodiffusion assay overestimates the C1q in serum. Here we describe a convenient, accurate procedure for measuring C1q in 250 microL of dialyzed serum. This method is based on our previous findings that all C1q in serum precipitates with the euglobulin fraction and that all other serum proteins containing hydroxyproline are excluded from this fraction. Because C1q is 4.3% hydroxyproline, the concentration of C1q in serum can therefore be calculated from the hydroxyproline content of the euglobulin fraction. The procedure, all done in the same tube, consists of precipitating the euglobulin fraction, digesting it with HClO4, and converting hydroxyproline to the corresponding pyrrole, which is extracted with toluene and measured by absorbance at 560 nm.
Exposure of streptomycin-resistant cells to puromycin results in uptake of dihydrostreptomycin comparable to that found with streptomycin-sensitive cells. This finding indicates that the enhanced phase of uptake, previously reported only in sensitive cells, may result from an increase in internal binding sites, presumably run-off ribosomes. The increased uptake of dihydrostreptomycin resulting from exposure to puromycin is greatest in both sensitive and resistant cells at concentrations below 100 microgram/ml. At 100 microgram/ml, exposure to puromycin in vivo results in significant, but not complete, polysome degradation and inhibition of protein synthesis. At 500 microgram/ml, where polysome degradation is complete in less than 2 min and where growth and protein synthesis are inhibited more than 90%, uptake of dihydrostreptomycin by both sensitive and resistant cells is inhibited. Puromycin has no effect on binding of dihydrostreptomycin to 70-S monosomes, as measured by equilibrium dialysis. The increased uptake of dihydrostreptomycin by resistant cells resulting from exposure to puromycin has no effect on viability. Addition of N-ethylmaleimide immediately and completely inhibits the puromycin-induced uptake of dihydrostreptomycin even when added after substantial polysome degradation has occurred.
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The binding of chloramphenicol to intracellular components of intact cells was measured by procedures based on a silicone-wash technique. The number of stereospecifically bound molecules of chloramphenicol increased with external concentration to a saturation value equal to the number of ribosomes per cell. Chloramphenicol is therefore believed to be attached stereospecifically by a weak bond, most probably to a single site on the 50S ribosome. This bond was found to be temperature-dependent and appeared to be responsible for inhibition of protein synthesis.
Hurwitz, Charles (Veterans Administration Hospital, Albany, N.Y.), Catherine B. Braun, and Richard A. Peabody. Washing bacteria by centrifugation through a water-immiscible layer of silicones. J. Bacteriol. 90:1692-1695. 1965.-A method is described which enables the separation of cells from aqueous suspension without altering the internal aqueous environment of the cells. The method consists of centrifuging the cells from the aqueous environment through a more dense, immiscible solvent consisting of a mixture of two silicones. A thin film of the aqueous environment equal to 7 x 10(-13) ml per cell remains with the bacteria during the separation procedure. The method by which this volume was determined is described. The procedure itself has no measurable effect on viability or permeability of the cells and permits recovery of about 90% of the cells from the aqueous environment. With this method, it has been found that the intracellular water volume of Escherichia coli ML 35 accessible to sucrose or inositol is 1.96 x 10(-12) ml +/- 6%, or 85% of the total volume of the cell determined by visual measurement.