Distinguishing of Ile/Leu amino acid residues in the PP3 protein by (hot) electron capture dissociation in Fourier transform ion cyclotron resonance mass spectrometry
Distinguishing of Ile/Leu amino acid residues in the PP3 protein by (hot) electron capture dissociation in Fourier transform ion cyclotron resonance mass spectrometry
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DOI：
10.1021/ac020422m
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发表时间：
2003-03-15
影响因子：
7.4
通讯作者：
Zubarev, RA
中科院分区：
文献类型：
--
作者：
Kjeldsen, F;Haselmann, KF;Zubarev, RA
在热电子捕获解离 (RECD) 中，捕获类似于 11-eV 的电子后会产生倍增质子化多肽片段。初级 c、z(.) 裂解时的能量过剩会导致 z(.) 片段发生二次断裂。由此产生的 w 离子使人们能够区分异构体 Ile 和 Len 残基。使用来自总共含有 135 个氨基酸残基的牛乳蛋白 PP3 的胰蛋白酶肽来评估 HECD 的分析效用。使用 Ile/Leu (Xle) 残基分配的正式程序，确定了 25 个 Xle 残基中的 20 个 (80%) 的身份。另外两个残基的身份可以根据替代 w 离子的不存在被正确猜测，并且只有两个残基（既没有观察到预期的也没有观察到替代 w 离子）仍然未分配。对传统 ECD 光谱的重新检查也揭示了 Xle w 离子的存在，尽管丰度较低，但所有 Xle 残基的 44% 都被识别出来。使用分配器阴极作为电子源，可以通过一次 2 秒长的采集来识别 2.7-kDa 肽中五个 Xle 残基中的四个，并通过对五次此类采集的平均值来识别所有五个残基。与高能碰撞诱导解离的情况不同，在胰蛋白酶肽的 HECD 中没有观察到 d 离子。
In hot electron capture dissociation (RECD), multiply protonated polypeptides fragment upon capturing similar to11-eV electrons. The excess of energy upon the primary c, z(.) cleavage induces secondary fragmentation in z(.) fragments. The resultant w ions allow one to distinguish between the isomeric Ile and Len residues. The analytical utility of HECD is evaluated using tryptic peptides from the bovine milk protein PP3 containing totally 135 amino acid residues. Using a formal procedure for Ile/Leu (Xle) residue assignment, the identities of 20 out of 25 Xle residues (80%) were determined. The identity of an additional two residues could be correctly guessed from the absence of the alternative w ions, and only two residues, for which neither expected nor alternative w ions were observed, remained unassigned. Reinspection of conventional ECD spectra also revealed the presence of Xle w ions, although at lower abundances, with 44% of all Xle residues distinguished. Using a dispenser cathode as an electron source, identification of four out of five Xle residues in a 2.7-kDa peptide was possible with one acquisition 2 s long, with identification of all five residues by averaging of five such acquisitions. Unlike the case of high-energy collision-induced dissociation, no d ions were observed in the HECD of tryptic peptides.