National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD, USA.
Rapid Commun Mass Spectrom. 2012 Apr 30;26(8):915-20. doi: 10.1002/rcm.6180.
Peptide identification reliability can be improved by excluding from analysis those m/z peaks of candidate peptides which cannot be observed in practice due to various physical, chemical or thermodynamic considerations. We propose using dissociation energies (as opposed to proton affinities) as a predictor of observability of different m/z peaks in spectra of short peptides.
Mass spectra of the tetrapeptides AAAA, AAFA, AAVA, AFAA, AVAA, AFFA, and AVVA were measured in the collision-induced dissociation (CID) activation mode on a grid of activation times 0.05 to 100 ms and normalized collision energy 10 to 35%. The lowest energy geometries and vibrational spectra were calculated for the precursor ions and their charged and neutral fragments using density functional theory (DFT) at the TPSS/6-31G(d,p) level. Dissociation energies were calculated for all fragmentation channels leading to b- or y-fragments.
It is demonstrated that m/z peaks observed in the mass spectra correspond to the fragmentation channels with the lowest dissociation energies. Using 50 kcal/mol as the cut-off value of dissociation energy, it was predicted that 28 out of 42 possible peaks in the b- and y-series of the seven tetrapeptides can be observed in mass spectra. In the experiments, 26 b- or y-peaks were observed, all of which are among the 28 predicted ones.
The use of dissociation energies generalizes the use of proton affinities for semi-quantitative predictions of relative intensities of different m/z peaks of short peptides. Further advances in this direction will pave the way for reliable quantitative predictions and, hence, for a significant improvement in robustness and accuracy of peptide and protein identification tools.
通过排除由于各种物理、化学或热力学考虑而在实践中无法观察到的候选肽的 m/z 峰，肽鉴定的可靠性可以得到提高。我们建议使用离解能（而不是质子亲和力）作为预测不同 m/z 峰在短肽光谱中可观察性的指标。
在碰撞诱导解离 (CID) 激活模式下，对 AAAA、AAFA、AAVA、AFAA、AVAA、AFFA 和 AVVA 这四种四肽的质谱进行了测量，在 0.05 到 100 ms 的激活时间网格和归一化碰撞能量 10 到 35%的条件下进行。使用密度泛函理论 (DFT) 在 TPSS/6-31G(d,p)水平上对前体离子及其带电和中性碎片的最低能量几何形状和振动光谱进行了计算。为所有导致 b-或 y-片段的碎裂通道计算了离解能。
证明了在质谱中观察到的 m/z 峰与具有最低离解能的碎裂通道相对应。使用 50 kcal/mol 作为离解能的截止值，预测出在这七种四肽的 b-和 y-系列中，42 个可能的峰中有 28 个可以在质谱中观察到。在实验中，观察到了 26 个 b-或 y-峰，它们都属于预测的 28 个峰中的峰。
离解能的使用扩展了质子亲和力的使用，可用于半定量预测短肽不同 m/z 峰的相对强度。这一方向的进一步发展将为可靠的定量预测铺平道路，从而为提高肽和蛋白质鉴定工具的稳健性和准确性提供显著的改进。
