Insight into the arginine biosynthetic pathway in Chlamydomonas by metabolite profiling

Christian Bölling1 and Lothar Willmitzer

Max Planck Institute for Molecular Plant Physiology, Am Mühlenberg 1, 14476 Potsdam, Germany; 1Current address: Charité Medical School, Monbijoustr. 2, 10117 Berlin, Germany

For most steps in arginine biosynthesis in Chlamydomonas mutants have been isolated that are deficient in the respective enzyme activities. This presents the unique opportunity to probe the potential of metabolite profiling for analyzing the effects of the introduction of specific breakpoints in a well-defined biochemical module through the loss of enzymatic activity. Three strains deficient in the activities of N-acetylglutamate-5-phosphate reductase (arg1), N2 acetylornithine-aminotransferase (arg9), and argininosuccinate lyase (arg2), respectively, were analyzed with regard to activation of endogenous arginine biosynthesis after withdrawal of externally supplied arginine. The different genetic backgrounds led to highly distinct changes in the levels for the intermediates of arginine biosynthesis. Enzymatic blocks could be characterized by precursor accumulation, like the amassment of argininosuccinate in arg2 cells, and depletion of downstream intermediates, e.g. N2-acetylornithine, ornithine, and argininosuccinate depletion in arg9 cells. The unexpected finding of substantial levels of N-acetylornithine, citrulline, and argininosuccinate downstream the enzymatic block in arg1 cells provided an explanation for the residual growth of these cells in the absence of external arginine sources. Their presence, together with the unusual accumulation of N-acetylglutamate, the first committed intermediate in ornithine and arginine biosynthesis, in arg1 cells suggests that alternative pathways, possibly involving the activity of ornithine aminotransferase, may be active when the default reaction sequence to produce ornithine via acetylation of glutamate is disabled.

e-mail address of presenting author: christian.boelling@charite.de