Charge state dependent collision-induced dissociation of native and reduced porcine elastase.
The [M + 20H](20+)-[M + 12H](12+) charge states of native and reduced porcine elastase, a 25.9 kDa serine protease, were subjected to collisional activation in a quadrupole ion trap. For most charge states, ion parking was used to increase the number of parent ions over that yielded directly by electrospray. Ion-ion proton transfer reactions were used to reduce product ion charge states largely to +1 to simplify spectral interpretation. Both forms of the protein show charge state dependent fragmentation behavior. The native protein, which contains four disulfide linkages, shows almost no evidence for fragmentation within the regions of the protein linked by disulfide bonds. However, at the lowest charge states studied, evidence for cleavage of a least one of the disulfide bonds was evident in the appearance of a c-type ion. The highest charge states of native elastase showed several prominent cleavages C-terminal to valine residues. As the charge state decreased, however, preferential cleavages at acidic amino acid residues became important. The reduced form of the protein did not show particularly prominent cleavages at valine residues. However, many of the same preferential cleavages at acidic amino acid residues noted for the native protein were also observed in the same charge states of the reduced protein. The reduced protein also showed additional cleavages from regions of the protein that are ordinarily protected by disulfide linkages in the native form.
Hogan
Jason M
JM
Department of Chemistry, Purdue University, West Lafayette, Indiana 47907-2084, USA.
McLuckey
Scott A
SA
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D007477:Ions / Q000737:chemistry; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010196:Pancreatic Elastase / Q000737:chemistry*; D011522:Protons; D055672:Static Electricity; D013552:Swine
D007477:Ions; D011522:Protons; D010196:Pancreatic Elastase