Abstract
The molecular weight forms of kynurenine formamidase were studied both genetically and biochemically. Formamidase I (native molecular weight 60,000) was purified using (NH4)2SO4 and pH fractionation, DEAE-cellulose chromatography at two different pH's, hydroxylapatite chromatography, and Sephadex G-100 gel filtration. Its subunit molecular weight, as determined by SDS gel electrophoresis, is 34,000, indicating that formamidase I is a dimer. Its K m is 1.87×10−3 m. Its isoelectric point is pH 5.3. Its amino acid composition is reported. Formamidase II (native molecular weight 31,000) was partially purified using techniques similar to those above. Its K m is 2.31×10−3 m. The response of formamidase activity to change in gene dosage was measured in segmental aneuploids generated in the second, third, and X chromosomes. Two separate chromosomal regions were identified which when present in extra dosage result in an elevation of the level of formamidase activity close to that predicted for the addition of a structural gene in a two-gene system. These tentative map positions were substantiated by demonstration that addition of one of the regions, 25A–27E, causes a 50% elevation in the relative amount of formamidase II. Addition of the other region, 91B–93F, causes a similar elevation in the relative amount of formamidase I. A model of the evolutionary origin of the two forms is presented, and the significance of these results to this model is discussed.
Similar content being viewed by others
References
Baily, C., and Wagner, C. (1974). Kynurenine formamidase purification and characterization of the adult chicken liver enzyme and chemical analysis of the enzyme of developing chicks. J. Biol. Chem. 2494439.
Cochran, D. (1976). Kynurenine formamidase in American cockroach. Insect Biochem. 6267.
Gabriel, O. (1971). Analytical disc gel electrophoresis. Methods Enzymol. 22565.
Glassman, E. (1956). Kynurenine formamidase in mutants of Drosophila. Genetics 41566.
Hall, J., and Kankel, D. R. (1976). Genetics of acetylcholinesterase in D. melanogaster. Genetics 83517.
Hodgetts, R. (1975). The response of dopa decarboxylase activity to variations in gene dose in D. melanogaster. Genetics 7945.
Kitos, B., and Sullivan, D. T. (1976). Developmental regulation of tryptophan catabolism in Drosophila. Insect Biochem. 6649.
Laemmli, O. K. (1970). Cleavage structure proteins during assembly of the head bacteriophage T4. Nature 227680.
Lindsley, D., Sandler, L., Baker, B., Carpenter, A., Denell, R., Hall, J., Jacobs, P., Miklos, G., Davis, B., Gethmann, R., Haroy, R., Hessler, A., Miller, S., Nozawa, H., Parry, D., and Gould-Somero, M. (1972). Segmental aneuploids and the genetic gross structure of the Drosophila genome. Genetics 71157.
Moore, G., and Sullivan, D. T. (1975). The characterization of multiple forms of kynurenine formamidase in D. melanogaster. Biochem. Biophys. Acta 397468.
O'Brien, S., and Gethman, R. (1973). Segmental aneuploids as a probe for structural genes in Drosophila: Mitochondrial membrane enzymes. Genetics 75155.
Shinohara, R., and Ishiguro, I. (1970). The purification and properties of formamidase from rat liver. Biochem. Biophys. Acta 198324.
Stewart, B. R., and Merriam, J. R. (1974). Segmental aneuploidy and enzyme activity as a method for cytogenetic localization in Drosophila melanogaster. Genetics 76301.
Sullivan, D. T., Kitos, R., and Grillo, S. (1974). Subcellular localization of the first three enzymes of the ommochrome synthetic pathway in Drosophila melanogaster. J. Exp. Zool. 188225.
Author information
Authors and Affiliations
Additional information
This work was supported by USPHS Grant GM-21286.
Rights and permissions
About this article
Cite this article
Moore, G.P., Sullivan, D.T. Biochemical and genetic characterization of kynurenine formamidase from Drosophila melanogaster . Biochem Genet 16, 619–634 (1978). https://doi.org/10.1007/BF00484718
Received:
Revised:
Issue Date:
DOI: https://doi.org/10.1007/BF00484718