Some tumor cells have a nearly absolute requirement for exogeneously supplied asparagine, while others can fulfill their requirements by endogenous biosynthesis. For this reason the enzyme responsible for the breakdown of asparagine has been used as a chemotherapeutic agent. The administration of inhibitors of the asparagine biosynthesis mechanism has also met with some therapeutic success. To enhance the effectiveness of selective tumor cell destruction, a complete understanding of asparagine biosynthesis is necessary. Much in fact remains unknown concerning the biosynthesis of asparagine. This ranges from determining the factors that control the asparagine synthesis rate in vivo to describing the metabolic needs for asparagine. We have recently shown that asparagine synthetase from both pancreas and liver can exist in three molecular forms; a 113,000, alpha-form, 57,000, beta-form and a 26,000 gamma-form. A dietary deficiency in asparagine results in the hepatic asparagine synthetase being in the alpha- the beta-forms. When animals are given dietary asparagine, only beta-asparagine synthetase is found. Treatment of animals with asparaginase also results in the appearance of the alpha-form of asparagine synthetase. We have shown that the crude beta-form can be converted to the alpha-form by incubation with glutamine and magnesium. We propose to examine the three different forms of asparagine synthetase and to establish their role in the control of asparagine metabolism in both normal and tumor cells. We have begun to explore the possible metabolic roles of asparagine in normal and tumor cell metabolism. We have found an enzyme in liver that can produce glyoxylate from isocitrate, and also another enzyme that uses asparagine and glyoxylate to form glycine. Possible links between asparagine and glycine metabolism are being investigated. Finding such a metabolic link may explain the selective cytotoxic effect exploited by asparaginase chemotherapy. It is hoped that these studies will uncover new targets for a rational approach to tumor chemotherapy.