Joseph F. Nyc

5 Glutamate Dehydrogenases

Publisher Summary This chapter discusses the types of investigations with major emphasis on recent studies that include elucidation of complete or partial amino acid sequences of the enzymes from several sources. L-Glutamate dehydrogenases catalyze the interconversion of α-ketoglutarate and L-glutamic acid. The enzymes are recognized to be important because of the pivotal positions in metabolism occupied by glutamate and α -ketoglutarate and the ability of these compounds to enter into many types of pathways. Glutamate dehydrogenases provide a route for incorporation of nitrogen into organic compounds, and thus a link between carbohydrate and amino acid metabolism. There are at least three types of glutamate dehydrogenases which differ in coenzyme specificity. Those specific for either NAD or NADP and those that can function with both. The successful isolation from some species of modified forms of the enzyme produced by mutant strains, particularly of Neurospora, now permits identification of residues important for maintenance of normal activity.

Properties of a phosphatidylmonomethylethanolamine N-methyltransferase from Neurospora crassa

Abstract A soluble preparation was obtained from microsomes of Neurospora crassa that catalyzed the two-step methylation of phosphatidylmonomethylethanolamine to phosphatidylcholine with S -adenosylmethionine as the methyl donor. Both the phosphatidyl- N -monomethylethanolamine and the phosphatidyl- N , N -dimethylethanolamine N -methyltransferase activities had optima near pH 8.0 and showed similar heat denaturation characteristics. The ratio of the two activities remained essentially the same in fractions obtained by precipitation with (NH 4 ) 2 SO 4 . The results of these studies are consistent with the conjecture based on previous work that one enzyme catalyzes the methylation of both phosphatidyl- N -monomethylethanolamine and phosphatidyl- N , N -dimethylethanolamine.

Chromatographic separation of estrone, estradiol and estriol.

Summary A column of pulverized Mealorub rubber has been adapted successfully to the quantitatively accurate chromatographic separation of a mixture of crystalline estrone, estradiol and estriol. When 20 ml each of 20, 40 and 60% aqueous methanol (v/v) are passed successively through the column, estriol is eluted by the 20%; estradiol by the 40% and estrone by the 60% concentration of methanol.

The accumulation of dimethylethanolamine by a mutant strain of Neurospora crassa

Abstract Dimethylethanolamine has been isolated from a mutant strain of Neurospor crassa . The chromatographic position of the isolated compound corresponds to that of authentic dimethylethanolamine. A cholineless mutant of Neurospora which requires either monomethylethanolamine, dimethylethanolamine, or choline for growth is able to grow in the presence of the isolated compound, and the growth response curve using increasing aliquots of the isolated dimethylethanolamine fraction follows the growth response curve observed with the authentic material. Carrier was added to the dimethylethanolamine isolated from a culture grown on 14 C-formate. The picric acid salt of the diluted radioactive dimethylethanolamine retained constantisotopic activity after repeated recrystallizations. This is the first time that dimethylethanolamine, a postulated precursor of choline, has been isolated from a natural source. The significance of the data has been discussed.

The chromatography of lipids in test tubes coated with a thin layer of silicic acid.

Abstract Test tubes, coated on the inner surface with a thin layer of silicic acid, are employed as the stationary phase for the ascending chromatography of various lipids. The chromatographic behavior of a number of purified lipids has been determined with mobile phases based on mixtures of chloroform with methyl alcohol and n-hexane with ethyl ether. The use of this system for the chromatography of lipid mixtures is illustrated with lipids extracted from different strains of Neurospora crassa.

The growth of niacin-requiring strains ofNeurospora crassa with normal and altered phospholipid compositions

Abstract The growth response of niacin-requiring strains ofNeurospora crassa to niacin supplements was employed as a means for estimating the metabolic availability of this vitamin to cells with normal and abnormal phospholipid compositions. A lecithin-deficient strain utilizes exogenous nicotinic acid and nicotinamide less efficiently for growth than does an organism with normal phospholipids. This poor growth response to niacin is more pronounced in surface-growing cultures than in liquid cultures. The growth of the lecithin-deficient mold on exogenous nicotinic acid is more dependent on the pH of the culture medium than is a lecithin-containing strain. Both the entrance of niacin into cells as well as the vitamins subsequent utilization for growth appear to be dependent on the phospholipid composition of the organism.

A colorimetric method for evaluating chymotrypsin inhibitors in human serum.

Summary A colorimetric method of determining serum chymotrypsin inhibitor levels has been descrived. This method has the following advantages over the original milk method described by West and Hilliard. 1 It is based on the hydrolysis of a stable, synthetic substrate. 2 Colorimetric determinations on timed reactions have obviated the need for subjective end point determinations.