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Abstract
Isotopic enrichment has diverse applications in MS for quantification, metabolite and peptide identification, metabolic tracing, and tandem mass tags to trigger MS2 events during data acquisition. Isotopic enrichment is also an important step for obtaining quality heteronuclear NMR data to measure the structure of intact proteins. High enrichment is desirable because the efficiency of the experiment scales as the product of the isotopic incorporation of the two elements, so it is desirable to perform quality control measurements to ensure complete enrichment. MS can be used to measure the percent incorporation of both 15N and 13C in uniformly enriched proteins at the intact level and the peptide level by comparing experimental data to many simulated isotope patterns with modified abundances of 15N and 13C. MS can also determine the site-specific incorporation of 15N and 13C in sparse isotope labeled proteins and measure the incorporation of labels in off-target sites due to metabolic scrambling. If more than one labeling composition is possible, relative isotopic mass defect can be used to discriminate between each possibility. Ultrahigh resolution MS measurements can reveal isotopic fine structure which can directly resolve each labeling possibility.