tailieunhanh - Báo cáo khoa học: Analysis of proteins and peptides on a chromatographic timescale by electron-transfer dissociation MS

Peptide and protein sequence analysis using a combination of gas-phase ion–ion chemistry and tandem MS is described. Samples are converted to multiply charged ions by ESI and then allowed to react with fluoranthene radical anions in a quadrupole linear ion trap mass spectrometer. | ỊFEBS Journal MINIREVIEW Analysis of proteins and peptides on a chromatographic timescale by electron-transfer dissociation MS Namrata D. Udeshi1 Jeffrey Shabanowitz1 Donald F. Hunt1 2 and Kristie L. Rose1 1 Department of Chemistry University of Virginia Charlottesville VA USA 2 Department of Pathology University of Virginia Charlottesville VA USA Keywords cell migration chromatin HIV regulator of expression of virion products mass spectrometry O-GlcNAcylation phosphorylation post-translational modifications protein identification Correspondence K. L. Rose Department of Chemistry University of Virginia Charlottesville VA 22904 USA Fax 434 982 2781 Tel 434 924 7994 E-mail klr6u@ Peptide and protein sequence analysis using a combination of gas-phase ion-ion chemistry and tandem MS is described. Samples are converted to multiply charged ions by ESI and then allowed to react with fluoranthene radical anions in a quadrupole linear ion trap mass spectrometer. Electron transfer from the radical anion to the multiply charged peptide or protein promotes random fragmentation along the amide backbone that is independent of peptide or protein size sequence or the presence of post-translational modifications. Examples are provided that demonstrate the utility of electron-transfer dissociation for characterizing post-translational modifications and for identifying proteins in mixtures on a chromatographic timescale 500 ms protein . Received 10 July 2007 revised 13 August 2007 accepted 17 October 2007 doi Introduction The traditional method of identifying proteins in complex mixtures by tandem MS involves the following steps a enzymatic digestion with trypsin b fractionation of the resulting tryptic peptides usually 10-30 residues in length by nanoflow HPLC interfaced to a mass spectrometer equipped for ESI c fragmentation of individual peptides by collision-activated dissociation CAD and d a search of the resulting tandem mass spectra