Shaw Jared B, Li Wenzong, Holden Dustin D, Zhang Yan, Griep-Raming Jens, Fellers Ryan T, Early Bryan P, Thomas Paul M, Kelleher Neil L, Brodbelt Jennifer S
Department of Chemistry and Biochemistry, The University of Texas at Austin, 1 University Station A5300, Austin, Texas 78712, USA.
J Am Chem Soc. 2013 Aug 28;135(34):12646-51. doi: 10.1021/ja4029654. Epub 2013 Jun 4.
The top-down approach to proteomics offers compelling advantages due to the potential to provide complete characterization of protein sequence and post-translational modifications. Here we describe the implementation of 193 nm ultraviolet photodissociation (UVPD) in an Orbitrap mass spectrometer for characterization of intact proteins. Near-complete fragmentation of proteins up to 29 kDa is achieved with UVPD including the unambiguous localization of a single residue mutation and several protein modifications on Pin1 (Q13526), a protein implicated in the development of Alzheimer's disease and in cancer pathogenesis. The 5 ns, high-energy activation afforded by UVPD exhibits far less precursor ion-charge state dependence than conventional collision- and electron-based dissociation methods.
由于蛋白质组学的自上而下方法有潜力提供蛋白质序列和翻译后修饰的完整表征，因此具有显著优势。在此，我们描述了在轨道阱质谱仪中实施193 nm紫外光解离（UVPD）以表征完整蛋白质的过程。通过UVPD可实现高达29 kDa的蛋白质近乎完全碎片化，包括对单个残基突变的明确定位以及对Pin1（Q13526）上几种蛋白质修饰的定位，Pin1是一种与阿尔茨海默病发展和癌症发病机制相关的蛋白质。UVPD提供的5 ns高能激活表现出比传统基于碰撞和电子的解离方法远更少的前体离子电荷态依赖性。
