AN ACCURATE MASS TAG STRATEGY FOR QUANTITATIVE AND HIGH THROUGHPUT PROTEOME MEASUREMENTS

ABSTRACT

We review a global strategy aimed at extending the breadth and throughput of proteomic measurements based on utilizing polypeptide “accurate mass tags” produced by global protein enzymatic digestion. The two-stage strategy described employs conventional mass spectrometric techniques to provide initial putative identifications of peptides based on searching genome databases using the partial sequence information gleaned from tandem mass spectrometric data. The second stage of the strategy utilizes advanced mass spectrometric instrumentation to “validate” those putative identifications initially made using conventional tandem mass spectrometric data. Specifically, the high mass accuracy provided by more advanced instrumentation is utilized and ultimately correlated with the conventional tandem mass spectrometric data (along with each of the peptides' liquid chromatographic elution time) to validate the putative identifications. The strategy described provides greater confidence in the assignment of protein identifications than possible with conventional tandem mass spectrometric data alone. The result is the generation of a database of accurate mass tags whose constituent peptides provide the basis for unambiguous identification of their cognate parent proteins and provide the basis for subsequent high throughput quantitative proteome measurements without the need for repeated tandem mass spectrometry.

ACKNOWLEDGMENTS

The authors wish to acknowledge the laboratory group of Dr. Richard D. Smith at Pacific Northwest National Laboratories.

This project has been funded in whole or in part with Federal funds from the National Cancer Institute, National Institutes of Health, under Contract No. NO1-CO-12400.

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