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, 2005 (2), 198-214

Metabolite Fingerprinting in Transgenic Nicotiana Tabacum Altered by the Escherichia Coli Glutamate Dehydrogenase Gene

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Metabolite Fingerprinting in Transgenic Nicotiana Tabacum Altered by the Escherichia Coli Glutamate Dehydrogenase Gene

R Mungur et al. J Biomed Biotechnol.

Abstract

With about 200,000 phytochemicals in existence, identifying those of biomedical significance is a mammoth task. In the postgenomic era, relating metabolite fingerprints, abundances, and profiles to genotype is also a large task. Ion analysis using Fourier transformed ion cyclotron resonance mass spectrometry (FT-ICR-MS) may provide a high-throughput approach to measure genotype dependency of the inferred metabolome if reproducible techniques can be established. Ion profile inferred metabolite fingerprints are coproducts. We used FT-ICR-MS-derived ion analysis to examine gdhA (glutamate dehydrogenase (GDH; EC 1.4.1.1)) transgenic Nicotiana tabacum (tobacco) carrying out altered glutamate, amino acid, and carbon metabolisms, that fundamentally alter plant productivity. Cause and effect between gdhA expression, glutamate metabolism, and plant phenotypes was analyzed by (13) NH(4)(+) labeling of amino acid fractions, and by FT-ICR-MS analysis of metabolites. The gdhA transgenic plants increased (13)N labeling of glutamate and glutamine significantly. FT-ICR-MS detected 2,012 ions reproducible in 2 to 4 ionization protocols. There were 283 ions in roots and 98 ions in leaves that appeared to significantly change abundance due to the measured GDH activity. About 58% percent of ions could not be used to infer a corresponding metabolite. From the 42% of ions that inferred known metabolites we found that certain amino acids, organic acids, and sugars increased and some fatty acids decreased. The transgene caused increased ammonium assimilation and detectable ion variation. Thirty-two compounds with biomedical significance were altered in abundance by GDH including 9 known carcinogens and 14 potential drugs. Therefore, the GDH transgene may lead to new uses for crops like tobacco.

Figures

Figure 1
Figure 1
Distribution of metabolites (judged by mass) altered in relative abundance in leaves and roots. Grey diamonds are leaf metabolites and black triangles represent metabolites altered in roots.
Figure 2
Figure 2
Scatter plot distribution of all classes of metabolites identified in leaf extracts.
Figure 3
Figure 3
Scatter plot distribution of all classes of metabolites identified in root extracts.
Figure 4
Figure 4
Metabolites in blue boxes were not detected. Metabolites in red boxes were used as internal standards and therefore detected. Metabolites in black boxes were detected and not changed. (a) Amino acids in leaves increased by gdhA. (b) Amino acids in roots increased by gdhA. Metabolites that are not protein amino acids are not annotated for changes here.
Figure 4
Figure 4
Metabolites in blue boxes were not detected. Metabolites in red boxes were used as internal standards and therefore detected. Metabolites in black boxes were detected and not changed. (a) Amino acids in leaves increased by gdhA. (b) Amino acids in roots increased by gdhA. Metabolites that are not protein amino acids are not annotated for changes here.

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