No abstract

The inactivation of aspartate aminotransferase in the course of the incubation with L-serine 0-sulfate is protected by thiosulfate. It has been found that also L-cysteine sulfinic acid is an inactivator when it is incubated with the enzyme, while L-serine-0-phosphate and L-cysteine-8-sulfonate are inactive. The inactivation by L-cysteine sulfinic acid is increa.sed by the simultaneous addition of 2-oxoglutarate and is protected by thiosulfate. Cysteine-8-sulfonate has been detected in the incubates of the enzyme with L-serine-0-sulfate protected with thiosulfate. It has been concluded that the protection by thiosulfate is due to the B-addition of thiosulfate to the inactivator aminoacrylic acid. The resulting cysteine-S-sdfonate thus represents an inactive dead-end product formed in the course of the protection by thiosulfate. It is suggested that this reaction could play the role of protecting enzymes sensitive to aminoacrylic acid and of inserting inorganic sulfur into aminoacrylic acid, producing a cysteine derivative. It has been recently reported [l] that pig heartaspartate aminotransferase is inactivated when it is incubated with L-serine-0-sulfate. The inactivation has been attributed to the production of aminoacrylic acid by the enzymatic @-elimination of the 0-sulfate group. Aminoacrylic acid is actually a strong reacting compound which adds spontaneously to a number of chemical groups before being cleaved into ammonia and pyruvic acid [2,3]. I n the attempt to use the enzymatic @-elimination of L-serine-0-sulfate by aspartate aminotransferase as a continuous source of aminoacrylic acid, to be used for other reactions, we have observed a protective effect exhibited by thiosulfate upon the inactivation of aspartate aminotransferase by L-serine-0-sulfate. The present paper describes this finding and a similar effect observed when L-cysteine sulfinic acid is used as an aminoacrylic-acid producer.

Pantothenoylcysteine-4'-phosphate decarboxylase has been purified 700-fold from horse liver by by an improved method. Enzyme activity was determined by measuring the amount of labeled C 0 2 produced from the labeled cysteine moiety of the substrate molecule. The enzyme is not bound to any cellular structure since more than 80 % of the total activity of the homogenate is recovered in the supernatant fraction. The enzyme does not seem to contain pyridoxal phosphate, is strongly inhibited by this latter and by carbonyl reagents, such as sodium borohydride, hydroxylamine and phenylhydrazine. These findings strongly support the presence in the enzyme molecule of a reactive carbonyl group. The enzymic reaction rate is increased over 20-fold by sulfhydryl compounds, has an absolute specificity for pantothenoylcysteine 4'-phosphate and is not inhibited either by pantothenoylcysteine or by the reaction product, pantetheine 4'-phosphate.

Screening by different means has demonstrated the presence, in human and murine neuroblastoma cell lines, of VGF, a gene product identified in a limited number of neuronal and endocrine cells. Indirect immunofluorescence and Western and Northern blot analyses have shown the presence of this protein in some of the tested lines, confirming that VGF is not an ubiquitous molecule. Further studies, using human SK-N-BE and murine N18TG2 lines, showed that VGF expression is upregulated during differentiation, suggesting that various species, including man, express VGF and regulate it in a similar manner. The subcellular localization of the protein, which is associated with vesicles, its electrophoretic molecular profile and its specific release under different conditions are all consistent with results reported in other cells. Neuroblastomas are thus added to the class of VGF-positive cells and provide a new in vitro model for investigation of the structural and functional properties of this protein.

Ex vivo rat brain microvessels express receptors for native as well as for oxidized low-density lipoproteins. In brain microvessels-derived endothelial cells, the expression levels of both receptors were enhanced by co-cultivation with rat astrocytes, even in the absence of actual contact between the two cell types, suggesting a soluble factor(s)-based mechanism of induction. No modulation e¡ect could be evidenced in a heterologous cellular system. Since both receptors were found to be expressed also in astrocytes, these cells are likely to contribute substantially to the lipoprotein management at the blood^brain barrier and in the brain compartment. ß