Identification of Rat Liver Aldehyde Oxidase across Species by Accurate Peptide-Mass Fingerprinting and Sequence-Tagging with Fourier Transform Ion Cyclotron Resonance Mass Spectrometry.

Abstract

We evaluated applicability of Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) to protein identification across species. Nano-electrospray FT-ICR MS enabled acquisition of accurate mass spectra of protein digests at a picomole level sensitivity. Peptide mass maps with a ppm level accuracy were routinely obtained by spraying whole protein digests without internal mass calibrant. The accurate peptide mass maps allowed narrowing of database search-windows for peptide-mass fingerprinting and the search noise was substantially reduced. This method of protein identification was applied to a rat liver protein, which had not been sequenced previously but had been biochemically characterized as an aldehyde oxidase. The rat liver protein was successfully correlated to human and bovine counterparts across the species boundaries by peptide-mass fingerprinting with an accurate tryptic mass map. Identity of the rat protein was further confirmed by accurate fragment-ion masses obtained by hexapole-storage-assisted capillary-skimmer dissociation of a peptide mixture.