Modulation of Protein Fragmentation Through Carbamylation of Primary Amines.
Modulation of Protein Fragmentation Through Carbamylation of Primary Amines.
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DOI：
10.1007/s13361-017-1648-5
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发表时间：
2017-08
影响因子：
3.2
通讯作者：
Brodbelt JS
中科院分区：
文献类型：
--
作者：
Greer SM;Holden DD;Fellers R;Kelleher NL;Brodbelt JS
我们评估了赖氨酸侧链和n端氨基氨基化对完整蛋白离子的断裂和大肠杆菌核糖体蛋白混合物的色谱特性的影响。比较了高能碰撞解离（HCD）和紫外光解离（UVPD）得到的6种未修饰和氨基甲酰化蛋白的断裂模式。由于碱性伯胺转化为非碱性氨基甲酸酯，氨基甲酰化显著减少了蛋白质保留的质子总数。氨甲酰化对HCD对蛋白质的片段化产生了显著的负面影响（即序列覆盖率降低和诊断片段离子减少），这与移动质子模型一致，该模型将肽片段化与电荷分布和电荷定向途径的机会联系起来。此外，氨基甲酰化蛋白在HCD上的N端和c端附近的碎片化增强。对于大肠杆菌核糖体蛋白的LCMS/MS分析，氨甲酰化后的保留时间平均增加了16分钟，这一结果归因于氨甲酰化后蛋白质的疏水性增加。对于这6种模型蛋白和核糖体蛋白，氨甲酰化对UVPD产生的片段离子的分布和类型的影响相对较小，这支持了完整蛋白的UVPD机制不反映移动质子模型的观点。
We evaluate the impact of carbamylation of the primary amines of the side-chains of Lys and the N-termini on the fragmentation of intact protein ions and the chromatographic properties of a mixture of E. coli ribosomal proteins. The fragmentation patterns of the six unmodified and carbamylated proteins obtained by higher energy collision dissociation (HCD) and ultraviolet photodissociation (UVPD) were compared. Carbamylation significantly reduced the total number of protons retained by the protein owing to the conversion of basic primary amines to non-basic carbamates. Carbamylation caused a significant negative impact on fragmentation of the protein by HCD (i.e., reduced sequence coverage and fewer diagnostic fragment ions) consistent with the mobile proton model, which correlates peptide fragmentation with charge distribution and the opportunity for charge-directed pathways. In addition, fragmentation was enhanced near the N- and C-termini upon HCD of carbamylated proteins. For LCMS/MS analysis of E. coli ribosomal proteins, the retention times increased by 16 min on average upon carbamylation, an outcome attributed to the increased hydrophobicity of the proteins after carbamylation. As noted for both the six model proteins and the ribosomal proteins, carbamylation had relatively little impact on the distribution or types of fragment ions product by UVPD, supporting the proposition that the mechanism of UVPD for intact proteins does not reflect the mobile proton model.
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影响因子：
7.4
作者：
Klein DR;Holden DD;Brodbelt JS
通讯作者：
Brodbelt JS
DOI：
10.1016/j.bbrc.2014.02.041
发表时间：
2014-03-21
影响因子：
3.1
作者：
Catherman, Adam D.;Skinner, Owen S.;Kelleher, Neil L.
通讯作者：
Kelleher, Neil L.
影响因子：
4.4
作者：
LeDuc, Richard D.;Fellers, Ryan T.;Early, Bryan P.;Greer, Joseph B.;Thomas, Paul M.;Kelleher, Neil L.
通讯作者：
Kelleher, Neil L.
影响因子：
7.4
作者：
Greer, Sylvester M.;Cannon, Joe R.;Brodbelt, Jennifer S.
通讯作者：
Brodbelt, Jennifer S.
影响因子：
7
作者：
Anderson, Lissa C.;Karch, Kelly R.;Hunt, Donald F.
通讯作者：
Hunt, Donald F.