tailieunhanh - Báo cáo khoa học: Top-down MS, a powerful complement to the high capabilities of proteolysis proteomics

For the characterization of protein sequences and post-translational modifi-cations by MS, the ‘top-down’ proteomics approach utilizes molecular and fragment ion mass data obtained by ionizing and dissociating a protein in the mass spectrometer. | ễFEBS Journal MINIREVIEW Top-down MS a powerful complement to the high capabilities of proteolysis proteomics Fred W. McLafferty1 Kathrin Breuker2 Mi Jin1 Xuemei Han1 Giuseppe Infusini1 Honghai Jiang1 z Xianglei Kong1 and Tadhg P. Begley1 1 Department of Chemistry and ChemicalBiology Baker Laboratory CornellUniversity Ithaca NY USA 2 Institute of Organic Chemistry and Center for Molecular Biosciences Innsbruck CMBI University of Innsbruck Austria Keywords electron capture dissociation MS protein characterization protein identification post-translationalmodifications top-down proteomics Correspondence F. W. McLafferty Baker Chemistry Laboratory CornellUniversity Ithaca NY 14853 USA Fax 607 255 4137 E-mail fwm5@ Received 30 May 2007 revised 12 October 2007 accepted 17 October 2007 doi For the characterization of protein sequences and post-translational modifications by MS the top-down proteomics approach utilizes molecular and fragment ion mass data obtained by ionizing and dissociating a protein in the mass spectrometer. This requires more complex instrumentation and methodology than the far more widely used bottom-up approach which instead uses such data of peptides from the protein s digestion but the topdown data are far more specific. The ESI MS spectrum of a 14 protein mixture provides full separation of its molecular ions for MS MS dissociation of the individual components. False-positive rates for the identification of proteins are far lower with the top-down approach and quantitation of multiply modified isomers is more efficient. Bottom-up proteolysis destroys the information on the size of the protein and the connectivities of the peptide fragments but it has no size limit for protein digestion. In contrast the top-down approach has a 500 residue 50 kDa limitation for the extensive molecular ion dissociation required. Basic studies indicate that this molecular ion intractability arises from greatly strengthened .

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