Absence of pyridoxine- (pyridoxamine-) 5'-phosphate oxidase in Morris hepatoma 7777

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Abstract
Morris hepatoma 7777 previously has been shown to have no detectable pyridoxine-(pyridoxamine-)5''-phosphate oxidase activity. To determine if this enzyme was missing in the hepatoma, rat liver oxidase was purified and antibodies to it were raised in rabbits. Final purification of rat liver oxidase for use as an antigen was accomplished by affinity chromatography and gel electrophoresis. The rat liver enzyme is similar to rabbit liver oxidase having 2 noncovalently linked subunits with MW in the range of 25,000-28,000. Evidence indicating that inactive enzyme was simultaneously purified with native enzyme was obtained. The IgG fraction was purified from the serum of a rabbit than had been immunized with rat liver oxidase. This was used in the development of enzyme-linked immunosorbent assay and immunoblot analyses for the presence of antigenically active pyridoxine- (pyridoxamine-) 5''-phosphate oxidase in cytosolic preparations from normal rat liver and Morris hepatoma 7777. There was no immunologically detectable oxidase protein in the tumor. An alternate pathway of pyridoxal 5''-phosphate synthesis, involving oxidation of pyridoxine to pyridoxal followed by phosphorylation, was ruled out. The implications of these findings with respect to acquisition of nutrients by tumors are discussed.

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