The Solubilization, Purification, and Properties of Nicotinamide Adenine Dinucleotide Glycohydrolase from Ehrlich Ascites Cells.

Treatment of mice bearing an Ehrlich ascites tumor with nitrogen mustard (HN2) results in an increase in the level of activity of the enzyme nicotinamide adenine dinucleotide glycohydrolase (EC 3.2.2.5) (1, 2). Comparison of some properties of this enzyme in homogenates of Ehrlich ascites cells from nitrogen mustard-treated and from untreated mice failed to reveal marked differences between the two. This finding suggested that the increase in activity in the former group might be due to an increase in the amount of enzyme protein (3). Experimental techniques used to demonstrate enzyme protein synthesis de novo generally employ partially purified soluble enzyme preparations (4, 5). With few exceptions NAD glycohydrolase has been found to be associated with the particulate fractions of cells (6, 7). Reported techniques for the solubilization of particulate NAD glycohydrolase vary, depending on the tissue source: extraction of erythrocyte stroma, pig brain, and beef spleen with isoamyl alcohol (8) ; digestion of pig spleen with trypsin (9) ; and extraction of an acetone powder of Aspergillus niger with 0.1 M potassium phosphate buffer, pH 7.5 (10). As a rule, appreciable losses in activity occur during these processes. Sslubilization of the NAD glycohydrolase, a particulate enzyme, from Ehrlich ascites cells (2) has not been reported. Attempts to apply previously reported methods to the solubilization of this enzyme from these tumor cells were unsuccessful. This paper presents a new technique for the solubilization of NAD glycohydrolase from Ehrlich ascites cells and from other mouse tissues. This procedure has been utilized in the purification of the enzyme from ascites cells. A comparison of the properties of the purified enzyme obtained from ascites cells of HN2-treated and from untreated mice is also presented.