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Characterization of Protein Carboxyl-O-methyltransferase in the Spontaneous in Vivo Murine C-1300 Neuroblastoma¹

Division of Clinical Pharmacology, Departments of Pediatrics and Pharmacology, University of Minnesota Medical School, Minneapolis, Minnesota 55455

Protein carboxyl-O-methyltransferase (PCM) activity was determined in subcellular fractions prepared from C-1300 neuroblastoma tumors following transplantation and growth in male A/J mice. Fractions were obtained by differential centrifugation, and PCM activity was determined in all fractions in the presence (+gel) and absence (-gel) of an exogenous substrate, gelatin. Sixty % of the PCM activity in the absence of exogenous substrate (-gel) was contained in the crude 800 x g particulate fraction, whereas 80% of the PCM activity in the presence of gelatin (+gel) was present in the postmicrosomal (100,000 x g) supernatant. The latter fraction also contained the highest specific activity of PCM. A K_m of 3.2 x 10^-6 M and a V_max of 5.3 pmol per mg protein per min were obtained for PCM activity (+gel) in the high-speed supernatant.

Cytoplasmic PCM was highly sensitive to competitive inhibition by S-adenosylhomocysteine and the S-adenosyl-homocysteine analogs sinefungin and A-9145C with K₁ values of 0.64, 0.47, and 0.05 µM, respectively. These data demonstrate that PCM present in murine neuroblastoma has characteristics similar to those of PCM isolated from other adrenergic and neuronal tissues. S-Adenosyl-homocysteine analogs may be useful probes for studying the role of PCM as a modulator of cell function in neurogenic and neoplastic tissues.

¹ Partially supported by grants from the Minnesota Medical Foundation, the Pharmaceutical Manufacturers Association Foundation, and USPHS (NS-17194). A preliminary report of this work was presented at the 65th Annual Meeting of the Federation of American Societies for Experimental Biology, Atlanta, Ga. (18).

² Research Career Development Awardee of the National Heart, Lung, and Blood Institute, USPHS (HL-00565). To whom requests for reprints should be addressed, at 3-260 Millard Hall, 435 Delaware St. S.E., Minneapolis, Minn. 55455.

³ Recipient of a Clinical Pharmacology postdoctoral fellowship from Ministere des Affaires Errangeres (France) and the Delegation Generale a la Recherche Scientifique et Technique (D. G. R. S. T., France).

Received 12/28/81. Accepted 7/26/82.