Protein identification in complex mixtures.
This paper investigates the prospects of successful mass spectrometric protein identification based on mass data from proteolytic digests of complex protein mixtures. Sets of proteolytic peptide masses representing various numbers of digested proteins in a mixture were generated in silico. In each set, different proteins were selected from a protein sequence collection and for each protein the sequence coverage was randomly selected within a particular regime (15-30% or 30-60%). We demonstrate that the Probity algorithm, which is characterized by an optimal tolerance for random interference, employed in an iterative procedure can correctly identify >95% of proteins at a desired significance level in mixtures composed of hundreds of yeast proteins under realistic mass spectrometric experimental constraints. By using a model of the distribution of protein abundance, we demonstrate that the very high efficiency of identification of protein mixtures that can be achieved by appropriate choices of informatics procedures is hampered by limitations of the mass spectrometric dynamic range. The results stress the desire to choose carefully experimental protocols for comprehensive proteome analysis, focusing on truly critical issues such as the dynamic range, which potentially limits the possibilities of identifying low abundance proteins.
Eriksson
Jan
J
Department of Chemistry, Swedish University of Agricultural Sciences, Box 7015, SE-750 07, Uppsala, Sweden. jan.eriksson@kemi.slu.se
Fenyö
David
D
D005656:Fungal Proteins / Q000737:chemistry*; D013058:Mass Spectrometry / Q000379:methods; D010455:Peptides / Q000302:isolation & purification
D005656:Fungal Proteins; D010455:Peptides