Jockusch R A, Schnier P D, Price W D, Strittmatter E F, Demirev P A, Williams E R
Department of Chemistry, University of California, Berkeley 94720, USA.
Anal Chem. 1997 Mar 15;69(6):1119-26. doi: 10.1021/ac960804q.
Blackbody infrared radiative dissociation spectra of the (M + 5H)5+ through (M + 11H)11+ ions of the protein ubiquitin (8.6 kDa) formed by electrospray ionization were measured in a Fourier-transform mass spectrometer. The 5+ ion dissociates exclusively by loss of water and/or ammonia, whereas the 11+ charge state dissociates only by formation of complementary y and b ions. These two processes are competitive for intermediate charge state ions, with the formation of y and b ions increasingly favored for the higher charge states. The y and b ions are formed by cleavage of the backbone amide bond on the C-terminal side of acidic residues exclusively, with cleavage adjacent to aspartic acid favored. Thermal unimolecular dissociation rate constants for the dissociation of each of these charge states were measured. From the temperature dependence of these rates, Arrhenius activation parameters in the rapid energy exchange limit are obtained. The activation energies (Ea) and preexponential factors (A) for the 5+, 8+, and 9+ ions are 1.2 eV and 10(12) s-1, respectively. These values for the 6+ and 7+ ions are 0.9-1.0 eV and 10(9) s-1, and those for the 10+ and 11+ ions are 1.6 eV and 10(16)-10(17) s-1. Thus, with the exception of the 5+ ion, the higher charge states of ubiquitin have larger dissociation activation energies than the lower charge states. The different A factors observed for production of y and b ions from different precursor charge states indicate that they are formed by different mechanisms, ranging from relatively complex rearrangements to direct bond cleavages. These results clearly demonstrate that the relative dissociation rates of large biomolecule ions by themselves are not necessarily a reliable indicator of their relative dissociation energies, even when similar fragment ions are formed.
在傅里叶变换质谱仪中测量了通过电喷雾电离形成的蛋白质泛素（8.6 kDa）的（M + 5H）5+至（M + 11H）11+离子的黑体红外辐射解离光谱。5+离子仅通过失去水和/或氨解离，而11+电荷态仅通过形成互补的y和b离子解离。这两个过程对于中间电荷态离子具有竞争性，对于较高电荷态，y和b离子的形成越来越受青睐。y和b离子仅通过酸性残基C端侧的主链酰胺键断裂形成，优先在天冬氨酸附近断裂。测量了这些电荷态中每种电荷态解离的热单分子解离速率常数。从这些速率的温度依赖性，获得了快速能量交换极限下的阿仑尼乌斯活化参数。5+、8+和9+离子的活化能（Ea）和指前因子（A）分别为1.2 eV和10(12) s-1。6+和7+离子的这些值为0.9 - 1.0 eV和10(9) s-1，10+和11+离子的值为1.6 eV和10(16)-10(17) s-1。因此，除了5+离子外，泛素的较高电荷态比较低电荷态具有更大的解离活化能。从不同前体电荷态产生y和b离子时观察到的不同A因子表明它们是通过不同机制形成的，范围从相对复杂的重排到直接键断裂。这些结果清楚地表明，即使形成相似的碎片离子，大生物分子离子自身的相对解离速率不一定是其相对解离能的可靠指标。
