The use of deoxyfluoro-d-galactopyranoses in a study of yeast galactokinase specificity

1. 2-Deoxy-2-fluoro-d-galactose, 3-deoxy-3-fluoro-d-galactose, 4-deoxy-4-fluoro-d-galactose, 6-deoxy-6-fluoro-d-galactose and 2-deoxy-d-lyxo-hexose are substrates for yeast galactokinase. 2. The variation in K(m) values for the d-hexose derivatives was not associated with a variation in the value of K(m) for MgATP(2-) indicating that the binding of MgATP(2-) is not modified by the binding of the sugar substrate. 3. Donated H bonds from OH-3, OH-4 and OH-6 and an accepted H bond to OH-2 of the d-hexose are important for the binding of the sugar substrate to galactokinase. 4. Yeast galactokinase exhibits similar kinetics to the galactokinase from Escherichia coli and operates by a similar random sequential mechanism. 5. 4-Deoxy-4-fluoro-d-glucose was neither a substrate for nor an inhibitor of yeast galactokinase.     

Water-soluble metabolites of p-nitrophenol and 1-naphthyl N-methylcarbamate in flies and grass grubs. Formation of glucose phosphate and phosphate conjugates

Metabolites isolated from houseflies dosed with 1-napththol or p-nitrophenol were identified as the phosphate and glucose phosphate conjugates of these phenols by titrations, hydrolysis, ionophoresis, i.r. spectra and mixed melting point. [(3)H]Carbaryl (1-naphthyl N-methylcarbamate) was metabolized by houseflies, blowflies and grass grubs to water-soluble metabolites which had chromatographic and ionophoretic behaviour similar to those of the conjugates of 1-naphthol with glucose, sulphate, phosphate and glucose 6-phosphate.     

Production and utilization of acetate in mammals

1. In an attempt to define the importance of acetate as a metabolic precursor, the activities of acetyl-CoA synthetase (EC 6.2.1.1) and acetyl-CoA hydrolase (Ec 3.1.2.1) were assayed in tissues from rats and sheep. In addition, the concentrations of acetate in blood and liver were measured, as well as the rates of acetate production by tissue slices and mitochondrial fractions of these tissues. 2. Acetyl-CoA synthetase occurs at high activities in heart and kidney cortex of both species as well as in rat liver and the sheep masseter muscle. The enzyme is mostly in the cytosol fraction of liver, whereas it is associated with the mitochondrial fraction in heart tissue. Both mitochondrial and cytosol activities have a K(m) for acetate of 0.3mm. Acetyl-CoA synthetase activity in liver was not altered by changes in diet, age or alloxan-diabetes. 3. Acetyl-CoA hydrolase is widely distributed in rat and sheep tissues, the highest activity being found in liver. Essentially all of the activity in liver and heart is localized in the mitochondrial fraction. Hepatic acetyl-CoA hydrolase activity is increased by starvation in rats and sheep and during the suckling period in young rats. 4. The concentrations of acetate in blood are decreased by starvation and increased by alloxan-diabetes in both species. The uptake of acetate by the sheep hind limb is proportional to the arterial concentration of acetate, except in alloxan-treated animals, where uptake is impaired. 5. Acetate is produced by liver and heart slices and also by heart mitochondrial fractions that are incubated with either pyruvate or palmitoyl-(-)-carnitine. Liver mitochondrial fractions do not form acetate from either substrate but instead convert acetate into acetoacetate. 6. We propose that acetate in the blood of rats or starved sheep is derived from the hydrolysis of acetyl-CoA. Release of acetate from tissues would occur under conditions when the function of the tricarboxylic acid cycle is restricted, so that the circulating acetate serves to redistribute oxidizable substrate throughout the body. This function is analogous to that served by ketone bodies.     

INTRACORPOREAL HEAT DISSIPATION FROM A RADIOISOTOPE-POWERED ARTIFICIAL HEART

The feasibility of radioisotope-fueled circulatory support systems depends on the ability of the body to dissipate the reject heat from the power source driving the blood pump as well as to tolerate chronic intracorporeal radiation. Our studies have focused on the use of the circulating blood as a heat sink. Initial in vivo heat transfer studies utilized straight tube heat exchangers (electrically and radioisotope energized) to replace a segment of the descending aorta. More recent studies have used a left ventricular assist pump as a blood-cooled heat exchanger. This approach minimizes trauma, does not increase the area of prosthetic interface with the blood, and minimizes system volume. Heat rejected from the thermal engine (vapor or gas cycle) is transported from the nuclear power source in the abdomen to the pump in the thoracic cavity via hydraulic lines. Adjacent tissue is protected from the fuel capsule temperature (900 to 1200 degrees F) by vacuum foil insulation and polyurethane foam. The in vivo thermal management problems have been studied using a simulated thermal system (STS) which approximates the heat rejection and thermal transport mechanisms of the nuclear circulatory support systems under development by NHLI. Electric heaters simulate the reject heat from the thermal engines. These studies have been essential in establishing the location, suspension, surgical procedures, and postoperative care for implanting prototype nuclear heart assist systems in calves. The pump has a thermal impedance of 0.12 degrees C/watt. Analysis of the STS data in terms of an electrical analog model implies a heat transfer coefficient of 4.7 x 10(-3) watt/cm(2) degrees C in the abdomen compared to a value of 14.9 x 10(-3) watt/cm(2) degrees C from the heat exchanger plenum into the diaphragm.     

Dihydrofolate reductase: low-resolution mass-spectrometric analysis of an elastase digest as a sequencing tool (Short Communication)

An elastase digest of a protein of unknown structure, dihydrofolate reductase, was studied by mass spectrometry. This soluble digest contained a large number of small peptides in different yields, within the ideal molecular-weight range (200-1200) for mixture-analysis mass spectrometry. Sequences of the major component peptides in the digest are reported.     

Transport of catecholamines by resealed chromaffin-granule `ghosts''

A method is described for the preparation of resealed chromaffin-granule ;ghosts'. The lysis and rapid purification procedures provide ;ghosts' in approximately 70% yield from crude granules; the preparation contains 0.1mumol of catecholamine/mg of protein (as compared with 2.8mumol/mg in unlysed granules), of which about one third is inside the ;ghosts'. The ;ghosts' retain their ability to accumulate catecholamines, a process dependent on Mg-ATP and inhibited by reserpine, and a simple assay for this transport is described.     

Concentrating Engines and the Kidney: II. Multisolute Central Core Systems

The analysis of the central core model of the renal medulla is extended to multisolute systems. It is shown that total solute concentration obeys the same differential equations for core and ascending limb as in a single solute system. Equations are derived for the concentration of individual solutes. Application of these equations to a two solute system shows that a central core system can concentrate with all transport being down a concentration gradient. This analysis applied to the renal medulla shows that mixing of urea from the collecting duct (CD) and salt from the loop of Henle in the central core of the inner medulla contributes to the concentration of urine during antidiuresis. It also sets criteria for completely passive function of the loop in the inner medulla, but whether these are satisfied cannot be determined from present experimental data.