Heterotropic interactions in monomeric βSH chains from human hemoglobin

The O₂ affinity of β^SH chains is lowered by H⁺, inositol hexaphosphate (IHP), and CO₂. As the oxygen affinity of β^SH monomers (β₁^SH) is lower than that of β^SH tetramers (β₄^SH), it is possible that IHP and CO₂ exert their influence on the O₂ affinity of β^SH chains by increasing the dissociation constant of β₄^SH rather than by a direct effect on the molecule. In order to test for this hypothesis we have measured the O₂ affinity of β^SH chains as a function of protein concentration at various concentrations of IHP and inorganic phosphates in the absence and presence of CO₂. From these data association constants for the binding of IHP to β₁^SH and β₄^SH as well as for the equilibrium 4β₁^SH ? β₄^SH were calculated. We found that IHP and CO₂ influence the oxygen affinity of β₁^SH. It was furthermore established that inorganic phosphate enhances the stability of β₄^SH while IHP favors its dissociation in monomers.     

Properties of a CDP-Diglyceride Hydrolase from Guinea Pig Brain

Enzymatic hydrolysis of the pyrophosphate bond of CDP-diglyceride (CDP-DG), previously shown to occur in bacteria, is demonstrable in mammalian tissues. Activity was enriched in a lysosomal fraction obtained from guinea pig cerebral cortex and was purified 92-fold relative to the homogenate by a combination of XM-300 ultrafiltration and DEAE-cellulose column chromatography. When incubated with CDP-dipalmitin, the purified enzyme produced stoichiometric amounts of CMP and phosphatidate. dCDP-DG served as a substrate, while ADP-DG was an inhibitor, as were 5'-AMP and 5'-dAMP. CDP-DG hydrolysis was not affected by the presence of excess amounts of CDP-choline, CDP-glycerol, sodium pyrophosphate, or cyclic 3',5'-AMP.     

Mammalian Brain Alkaline Phosphatase: Expression of Liver/Bone/Kidney Locus Comparison of Fetal and Adult Activities

The alkaline phosphatases (EC 3.1.3.1) are determined by at least three gene loci, which can be sharply distinguished one from another by their sensitivity to inhibition with various amino acids and peptides and by ther-mostability. Alkaline phosphatase is present in the brains of guinea pig, rat, mouse, hamster, squirrel, rabbit, cat, sheep, cow, tamarin, baboon, and man. The gene locus coding for alkaline phosphatase in all these brains is the liver/ bone/kidney locus, as indicated by thermostability studies and by inhibition studies with L-phenylalanine, L-homoarginine, and L-phenylalanylglycylglycine. The average brain alkaline phosphatase activity is about 35% of the average for the livers and only 7.2% and 4.4% of the average kidney and placental activities, respectively. During growth and development, brain alkaline phosphatase activity decreases in the mammals studied. The amount of change is tissue- and species-dependent.     

Development of cyanide-resistant respiration in mitochondria from potato tubers treated with ethanol,acetaldehyde, and acetic acid

Treatment of intact potato (Solanum tuberosum L.) tubers with acetaldehyde, ethanol or acetic-acid vapors led to a respiratory upsurge which was further increased when the volatiles were applied in 100% O₂. Mitochondria from tubers held in 100% O₂ (O₂ control) displayed a substrate state, state 3, and state 4 in respiration, whereas in mitochondria from the volatile-treated tubers the respiratory rate of the different states was virtually indistinguishable. This respiratory pattern was companied by the development of a cyanide-resistant respiration since these mitochondria exhibited resistance to CN and sensitivity to CN+salicylhydroxamic acid. Acetaldehyde-treated potatoes showed a time-course development (up to 36 h) of cyanide resistance and concomitant sensitivity to salicylhydroxamic acid, indicating the onset of synthetic processes leading to the observed changes in mitochondrial respiration.Abbreviations V total respiration rate - V_cyt velocity of O₂ uptake attributable to cytochrome oxidase - V_alt velocity of O₂ uptake attributable to the alternate oxidase - RCR respiratory control ratio - SHAM salicylhydroxamic acid Paper of the Journal Series, New Jersey Agricultural Experiment Station, Cook College, Rutgers University, New Brunswick, N.J., USA     

A Standardized Test of Renal Concentrating Capacity in Adults with Some Results in Essential Hypertension

Three groups of patients: A with normal glomerular filtration rate, B with moderate and C with advanced renal damage, were dehydrated and fasted for 30 hours. At regular intervals measurements were taken of urine osmolality, urine specific gravity and serum osmolality. The time required to reach maximum urine osmolality varies with the degree of dehydration and inversely with the severity of kidney damage. In patients with normal glomerular filtration rate, maximum urine osmolality is not attained by 30 hours of dehydration. Thus, for shorter periods, all “normal ranges” of concentrating capacity must be related to specific durations of dehydration. Carefully measured urine specific gravities parallel urine osmolalities closely, especially when proteinuria and glucosuria are absent. The measurement of U/P osmolality ratio offers no clinical advantage in the assessment of renal concentration capacity over the measurement of urine osmolality alone. In Group A, hypertensives achieved higher urine concentrations than did the nonhypertensives under identical test conditions. A normal range for renal concentrating capacity has been presented.     

Dicyclohexylammonium salts for the isolation and characterization of aldonic acids

Dicyclohexylammonium salts of aldonic acids may be prepared from aldonolactones, as well as from metal aldonates and the free acids. Although accompanied by decomposition, their melting points are usually sharp, and these salts appear to have some potential utility for the isolation and characterization of aldonic acids.Dicyclohexylammonium 2-acetamido-2-deoxy-d-gluconate (1) has recently been described; in the course of the present investigation, it was converted into 2-acetamido-2-deoxy-d-glucose (3), confirming the configuration previously assigned to it. With aqueous dicyclohexylamine, 2-acetamido-2-deoxy-d-mannono-1,4-lactone (2) gives 1. The configuration of 2 was reconfirmed through reduction to 2-acetamido-2-deoxy-d-mannitol (4), and the optical rotations of this compound and its d-gluco isomer in acidified ammonium molybdate solution were found to be useful physical constants for distinguishing these alditols.2-Acetamido-2-deoxy-d-galactono-1,4-lactone affords a crystalline dicyclohexylammonium salt of the corresponding acid, from which the lactone may be regenerated.     

Scanning isoelectric focusing in small density-gradient columns: I. Use of a Standard Spectrophotometer Cuvette for Focusing. Chemical Modification of Proteins by Migrating Reactive Ions

By the method of isoelectric focusing in a sucrose density gradient, small protein samples (less than 100 μg) have been separated and analyzed within 2 hr, using as electrolysis column a commercial standard quartz spectrophotometer cuvette, equipped with platinum electrodes and placed in an optical scanning device. Preparation of the cuvette prior to an isoelectric focusing experiment required about half an hour with no external apparatus, as the density gradient was created spontaneously in the cell by free interdiffusion of sucrose solutions. The cuvette temperature could be controlled by circulating water. The optical detection device permitted repeated scanning of the cuvette during the electrolysis process, thus providing information about the events occurring to a protein during focusing or prolonged electrolysis. By scanning with wavelength at the positions where the proteins have focused, their absorption spectra were obtained. the isoelectric points of separated proteins were estimated by fractionation of the cell contents and subsequent pH measurements on the fractions.The present paper also describes how individual Ampholine components, or groups of components, in their focused states gave rise to easily detected refractive-index gradients within the cell. The optical scanning device has been built in such a way that interference of these gradients with absorption measurements was abolished.Application of the technique to the isoelectric separation of commercial sperm whale myoglobin is reported. Ferrous or ferric forms of the focused myoglobin components were obtained by migration of reducing or oxidizing agents through the zones.     

Ultrastructural localization of monoamines in the central nervous system of Lumbricus terrestris (L.) with remarks on neurosecretory vesicles

Summary The cerebral ganglion and the ventral nerve cord of Lumbricus terrestris have been studied with the electron microscope. The results are as follows: In the neuropile small granular vesicles (300 to 500 Å) occur in some varicose nerve fibres after fixation with potassium permanganate. This indicates the presence of noradrenaline. Sometimes only a few of the vesicles produce a positive reaction. After incubation with -methyl-noradrenaline the numbers of nerve terminals with small granular vesicles greatly increase, indicating the presence of dopamine and/or 5-hydroxytryptamine. In this case the reaction is now complete. The number of small granular vesicles is largest in the terminal swellings.These findings are consistent with histofluorescence, chemical, and microspectrofluorometric analyses, which have demonstrated noradrenaline, dopamine, and 5-hydroxytryptamine in neurones in the central nervous system.Large granular vesicles (600 to 900 Å) are to be found in some perikarya, not identical with neurosecretory cell bodies. In this case the granular vesicles in the axon are smaller and fewer. This indicates a simultaneous proximo-distal transport and gradual decrease in size of the granular vesicles. The intraneuronal distribution of the vesicles is in agreement with the distribution of the fluorophores in the fluorescent neurones.Neurosecretory neurones are found most likely not to contain monoamines.This work was supported by grants from the Helge Axison Johnson Foundation, the Magn. Bergvall Foundation, and the Roy. Physiographic Society at Lund.I am greatly indebted to Mrs. Lena Eriksson, Miss Rita Jönsson, Miss Inger Norling, Mrs. Lena Svenre, and Mr. Henryk Keff for their excellent technical assistance.     

Contributions of various ions to the resting and action potentials of crayfish medial giant axons

Summary The membrane of crayfish medial giant axons is permeable at rest to ions in the rank K>Na>Ca>Cl. With K present, variation of the other ions has little or no effect, but with K absent the axon hyperpolarizes when Na is reduced or eliminated by replacement with Tris (slope ca. 30 mV/decade Na₀). The hyperpolarization is independent of the presence of Cl or its absence (substitution with methanesulfonate or isethionate). The resistance increases progressively as Na is removed. These changes persist after the spike is blocked with tetrodotoxin. An increase in Ca causes depolarization (slope ca. 20 mV/decade) provided K, Na and Cl are all absent, but in the presence of Cl there is little or no change in membrane potential on increasing Ca to 150mm. The depolarization induced by Ca is associated with an increased resistance. Spike electrogenesis involves Ca activation as well as Na activation, but the after-depolarization at the end of the spike is due to a conductance increase for Ca. Two alternative equivalent circuits for the resting and active membrane are discussed.     

BENTHIC ALGAL COMMUNITIES OF THE METOLIUS RIVER1,2

Sampling of benthic algae from selected sites in the Metolius River was carried out at regular intervals between April 15, 1967 and March 9, 1968 to determine the species composition of the communities. With the exception of the sampling site at the headwaters and the site immediately downstream, the benthic community of the Metolius River is described as a Cladophora–Achnanthes–Spirogyra community, with species of these 3 genera predominating on a year-round basis. The community described for the headwaters and the sampling site immediately downstream was composed primarily of Nitzchia palea and Cymbella cistula. Of the approximately 60 species of algae which were identified, only 9 showed a definite seasonal distribution, the remaining species being found on a year-round basis.