13C metabolic flux analysis is the technology of choice to experimentally determine metabolite exchange rates (fluxes) in central metabolism. Fluxes are determined by feeding 13C labeled substrates to cells, and measuring the label distribution in (mostly) amino acids using mass spectrometry and nuclear magnetic resonance.
Peptide-based 13C metabolic flux analysis (MFA) takes advantage of the peptide sequence to provide an identity for each isotope label measurement. Because each measurement has an identity, flux maps unique to the peptide origin can be established without the need to purify protein from a target cell-type, organelle or developmental stage. Tryptic digests of extracted protein from an entire organism can be analyzed on a high resolution pass spectrometer and yield tissue specific flux maps.
Of special interest to biotechnology is the application of peptide-based MFA to mutant libraries. Knockout libraries that are prepared with transposons expressing a set of peptides specific to each mutant, allow for the characterization of central metabolism of thousands of mutants with a single 13C labeling experiment.
To take full advantage of the large mass spectrometry datasets, labeled peptides need to be identified and quantified fully automatically. GenomeCraft can extract ~90% of observed unlabeled peptide measurements unsupervised, at a fidelity that exceeds that of a trained professional.
The CEO of GenomeCraft pioneered peptide-based flux analysis, and offers custom-tailored flux analysis ranging from conventional to highly novel approaches. Please contact GenomeCraft to explain your plans, and hear about the available options and associated cost.
Relevant publications