Genes of Nitrogen Metabolism in Escherichia Coli

University Of Texas At Dallas, Richardson TX

Investigators

Abstract

The working hypotheses of this research are that E. coli contains two putrescine catabo-lism pathways, and that both modulate intracellular polyamines, albeit during different growth conditions. The first pathway has been previously described, and involves two transaminase-dependent deaminations, and two dehydrogenase-dependent oxidations. The second pathway has not been previously described, and appears to involve glutamylated intermediates. To test these hypotheses, mutants lacking each of the proposed genes of putrescine catabolism will be con-structed, and the resulting phenotypes examined. Enzymes of the proposed pathways will be pu-rified and characterized. The functions of the two pathways will be tested by examination of the expression of putrescine catabolic genes, and assay of the activities of the gene products in wild type strains and in strains lacking specific regulators. Finally, intracellular polyamines will be extracted, separated, and quantified in order to test whether the proposed intermediates accumu-late in specific mutants, and whether these pathways modulate intracellular polyamines. This study will provide insight into a fundamental, but poorly understood, aspect of me-tabolism, and into the growth and metabolism of bacteria. Polyamines, such as putrescine and spermidine, are required for the rapid growth of essentially all organisms. The role of degrada-tive enzymes in the control of polyamine levels has not been examined because the degradative pathways have not been determined. This study will elucidate a degradative pathway that has not been described in any organism, and will determine the extent that the degradative pathways control polymaine levels. Undergraduates constructed a majority of the mutants that provided the unpublished background for this project. They will continue to be an integral part of the discov-ery process.