Elucidation of novel biosynthetic pathways and metabolite flux patterns byretrobiosynthetic NMR analysis

The labelling patterns of metabolites from experiments with stable isotope- labelled precursors can be determined by NMR spectroscopy. Complex isotopom er mixtures are found when general metabolites such as glucose are used as stable isotope-labelled precursors which are diverted to all branches of in termediary metabolism. The complex results can be interpreted by a pattern recognition approach based on comparison between the labelling patterns of secondary metabolites and primary metabolites such as amino acids and ribon ucleosides. The isotope labelling patterns of intermediates in central meta bolic pools such as carbohydrate phosphates, dicarboxylic acids, and acetyl CoA can be obtained by biosynthetic retroanalysis. Biosynthetic pathways a s well as metabolite flux patterns can be determined from these data. The m ethod is illustrated using the classical mevalonate pathway and the more re cently discovered deoxyxylulose pathway of terpenoid biosynthesis as exampl es. Applications of the retrobiosynthetic method of the biosynthesis of mol ybdopterin and of riboflavin are also discussed. Stable isotope experiments monitored by NMR spectroscopy have also been shown to be a powerful tool f or the elucidation of metabolic flux in microorganisms with unusual lifesty les and in fermentation processes. (C) 1999 Published by Elsevier Science B .V. All rights reserved.