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ASPARAGINE AND GLYCINE METABOLISM IN RAT LIVER MITOCHONDRIA AND IN MOUSE L5178Y LYMPHOMA CELLS RESISTANT OR SENSITIVE TO THE ANTICANCER DRUG L-ASPARAGINASE (LEUKEMIA, CITRIC ACID CYCLE, GLYOXYLATE, AMINOTRANSFERASE, SERINE)
Abstract
Rat liver mitochondrial asparagine was found to be degraded via an aminotransferase and (omega)-amidase. Evidence includes oxaloacetate production from asparagine only when glyoxylate was added and production of radiolabeled (alpha)-ketosuccinamate via metabolism of U-('14)C asparagine. In the cytosol, asparagine is degraded primarily via asparaginase and subsequent transamination. A new HPLC technique for separation of citric acid cycle intermediates was developed using: ion pairing with 20 mM each to tetrabutylammonium hydroxide and Na(,2)SO(,4); pH 7.0; isocratic elution; and detection at 210 nm. Amino acids were analyzed using precolumn derivitization with ortho-phthaldialdehyde. A computer program was written in BASIC to process amino acid analyses data and perform error analysis. Amino acid content of mouse lymphoma cells either sensitive (L5178Y) or resistant (L5178Y/L-ASE) to the anticancer drug L-asparaginase was studied. The concentration of asparagine was 1.5 times higher and the concentrations of the essential amion acids histidine, methionine, valine and phenylalanine were two times higher in asparaginase-resistant than sensitive cells. In vivo but not in vitro studies indicated that glycine decreases in sensitive but not resistant cells upon asparaginase treatment. Asparagine and glycine metabolism was further studied using ('14)C radiolabel conversion of asparagine, glyoxylate, glycine and serine. The conversion of glycine to serine in sensitive cells was 2.5 times higher than resistant cells. Upon incubation with asparaginase, glyoxylate to glycine conversion decreased 20% and serine to glycine conversion increased 79% in sensitive cells but did not change in resistant cells. These results indicate asparagine dependent differences in glycine metabolism between the two cell lines. Glycine metabolism is especially important in lymphomas and leukemias because these cells contain higher concentrations of glycine that other cancer and normal cells. Therefore, glycine levels were studied and were found to decrease in sensitive but not resistant cells upon asparaginase administration. Also partially purified glyoxylate-asparagine aminotransferase from L5178Y/L-ASE (resistant) cells was found to have a five fold higher apparent V(,max) than L5178Y (sensitive ) cell glyoxylate-asparagine aminotransferase. This may partially account for asparaginase resistance; the higher enzyme activity may prevent a lethal asparaginase-induced decrease in glycine concentration in resistant but not asparaginase sensitive cells.
Subject Area
Biochemistry
Recommended Citation
KEEFER, JAMES F., "ASPARAGINE AND GLYCINE METABOLISM IN RAT LIVER MITOCHONDRIA AND IN MOUSE L5178Y LYMPHOMA CELLS RESISTANT OR SENSITIVE TO THE ANTICANCER DRUG L-ASPARAGINASE (LEUKEMIA, CITRIC ACID CYCLE, GLYOXYLATE, AMINOTRANSFERASE, SERINE)" (1986). ETD collection for University of Nebraska-Lincoln. AAI8614456.
https://digitalcommons.unl.edu/dissertations/AAI8614456