PNNL

Abstract

We describe methods for mass spectrometric identification of heme-containing peptides from digests of c-type cytochromes that contain the CXXCH(X = any amino acid) sequence motif. Analysis of purified standard heme-containing peptides showed that the charged heme group was present both before and after peptide fragmentation in the gas phase. The heme fragment ion yielded the most abundant MS/MS peak for standard heme-containing peptides with one amino acid difference (DAA=1) for both 2+ and 3+ peptide charge states and the extent of heme loss during peptide fragmentation was affected by both sequence and charge. A modified search strategy was evaluated with tryptic digests of one known and two unknown cytochromes from Shewanella oneidenis, demonstrating that this approach can be generally applied for identification of c-type heme-containing peptides from complex samples.