Charge state dependent collision-induced dissociation of native and reduced porcine elastase
Charge state dependent collision-induced dissociation of native and reduced porcine elastase
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DOI：
10.1002/jms.458
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发表时间：
2003-03-01
影响因子：
2.3
通讯作者：
McLuckey, SA
中科院分区：
文献类型：
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作者：
Hogan, JM;McLuckey, SA
25.9 kDa丝氨酸蛋白酶天然和还原猪弹性蛋白酶的[M+20H](20+)-[M+12H]12(+)电荷态发生碰撞。四极离子的活化对于大多数电荷态，离子停滞被用来增加母体离子的数量，而不是直接通过电喷雾产生的。利用离子-离子质子转移反应将产物离子电荷态大大降低到+1，以简化光谱解释。这两种形式的蛋白质都表现出依赖于电荷状态的碎裂行为。这种天然蛋白质含有四个二硫键，几乎没有证据表明蛋白质中由二硫键连接的区域存在裂解。然而，在所研究的最低电荷态，至少一个二硫键断裂的证据在C型离子的出现中是明显的。天然弹性蛋白酶的最高电荷态显示出几个突出的裂解C-末端到Valine残基。然而，随着电荷状态的降低，酸性氨基酸残基上的优先裂解变得重要。还原形式的蛋白质在Valine残基上没有表现出特别明显的裂解。然而，许多与天然蛋白质相同的酸性氨基酸残基上的优先裂解也在还原蛋白质的相同电荷状态下观察到。还原的蛋白质还显示出来自蛋白质区域的额外裂解，这些区域通常由天然形式的二硫键保护。版权所有(C)2003 John Wiley Sons，Ltd.
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 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. Copyright (C) 2003 John Wiley Sons, Ltd.