Metabolic action of cortisone; D-amino acid oxidase and apparent creatinine formation

Data are presented demonstrating that the d-amino acid oxidase of rat liver is lowered on adrenalectomy and restored to normal by treatment with cortisone. This reaction is responsible for data obtained which indicated that creatine and creatinine formation were under the control of cortisone. The α-keto-γ-methiolbutyric acid formed from the oxidation of methione reacts in the Jaffe reaction as “creatinine.” During the course of incubation of the liver enzyme system a portion of the α-keto-γ-methiolbutyric acid appears in a bound form and is measured by way of the Jaffe reaction as “creatine.” Measurements of creatine formation by a method not subject to these errors show that creatine formation in the rat liver enzyme is very small although creatine formation in guinea pig liver can readily be demonstrated.     

Immune complex effects on murine macrophages. I. Immune complexes suppress interferon-gamma induction of Ia expression

We have studied the effects of immune complexes on the expression of macrophage surface proteins in vitro. Increased expression of the H-2 molecules I-A, I-E, and K on the macrophage membrane was induced by in vitro culture with crude lymphokine or interferon-gamma. Expression of all three of the molecules was additionally increased by stimulating the cultures with heat-killed Listeria monocytogenes. Addition of soluble immune complexes to the cultures did not have any effect on macrophage expression of these proteins. However, significant inhibition of lymphokine or interferon-gamma induction of I-A, I-E, and H-2K was observed when macrophages were cultured on plates to which immune complexes had been bound. This inhibition was dose dependent, required an immunoglobulin (Ig) molecule with an intact Fc portion, did not require the presence of T cells, and occurred in the presence of indomethacin. Complexes containing IgG1, IgG2a, IgG2b, and IgE, but not IgM or IgA, antibodies mediated the inhibitory effect.     

Measurement of positional isotope exchange rates in enzyme-catalyzed reactions by fast atom bombardment mass spectrometry: application to argininosuccinate synthetase

Fast atom bombardment mass spectrometry (FAB-MS) has been used to measure positional isotope exchange rates in enzyme-catalyzed reactions. The technique has been applied to the reactions catalyzed by acetyl-CoA synthetase and argininosuccinate synthetase. The FAB technique is also able to quantitatively determine the oxygen-18 or oxygen-17 content of nucleotides on as little as 10 nmol of material with no prior derivatization. Acetyl-CoA synthetase has been shown by FAB-MS to catalyze the positional exchange of an oxygen-18 of ATP from the beta-nonbridge position to the alpha beta-bridge position in the presence of acetate. These results are consistent with acetyl adenylate as a reactive intermediate in this reaction. Argininosuccinate synthetase was shown not to catalyze a positional isotope exchange reaction designed to test for the formation of citrulline adenylate as a reactive intermediate. Argininosuccinate synthetase was also found not to catalyze the transfer of oxygen-18 from [ureido-18O]citrulline to the alpha-phosphorus of ATP in the absence of added aspartate. This experiment was designed to test for the transient formation of carbodiimide as a reactive intermediate. These results suggest that either argininosuccinate synthetase does not catalyze the formation of citrulline adenylate or the enzyme is able to completely suppress the rotation of the phosphoryl groups of PPi.     

Possible regulation of pyruvate carboxylase fromThiobacillus novellus by hydroxypyruvate

Aspartate, glutamate, or dicarboxylic acids did not inhibit the activity of a highly purified but not homogeneous preparation of pyruvate carboxylase fromThiobacillus novellus. The only effective inhibitors were end-products of the reaction and to a lesser degree hydroxypyruvate. The latter has not been shown previously to regulate the enzyme's activity. Lineweaver-Burk plots revealed that it was uncompetitive with respect to acetyl CoA with a K_ii of 3.6 mM, and noncompetitive with respect to bicarbonate, magnesium ATP, and pyruvate with respective K_ii values of 7.1, 5.5, and 6.47 mM. The corresponding K_is values were 7.02, 5.4, and 4.25 mM. A mathematical model is presented that supports the findings.