Comprehensive Characterization of Swine Cardiac Troponin T Proteoforms by Top-Down Mass Spectrometry

doi:10.1007/s13361-018-1925-y

	Comprehensive Characterization of Swine Cardiac Troponin T Proteoforms by Top-Down Mass Spectrometry
	Lin, Ziqing 1,2; Guo, Fang 1,3; Gregorich, Zachery R.1; Sun, Ruixiang 1,4; Zhang, Han 5; Hu, Yang 1; Shanmuganayagam, Dhanansayan 6; Ge, Ying 1,2,5
	2018-06-01
发表期刊	JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY
ISSN	1044-0305
卷号	29 期号:6 页码:1284-1294
摘要	Cardiac troponin T (cTnT) regulates the Ca2+-mediated interaction between myosin thick filaments and actin thin filaments during cardiac contraction and relaxation. cTnT is released into the blood following injury, and increased serum levels of the protein are used clinically as a biomarker for myocardial infarction. Moreover, mutations in cTnT are causative in a number of familial cardiomyopathies. With the increasing use of large animal (swine) model to recapitulate human diseases, it is essential to characterize species-dependent protein sequence variants, alternative RNA splicing, and post-translational modifications (PTMs), but challenges remain due to the incomplete database and lack of validation of the predicted splicing isoforms. Herein, we integrated top-down mass spectrometry (MS) with online liquid chromatography (LC) and immunoaffinity purification to comprehensively characterize miniature swine cTnT proteoforms, including those arising from alternative RNA splicing and PTMs. A total of seven alternative splicing isoforms of cTnT were identified by LC/MS from swine left ventricular tissue, with each isoform containing un-phosphorylated and mono-phosphorylated proteoforms. The phosphorylation site was localized to Ser1 for the mono-phosphorylated proteoforms of cTnT1, 3, 4, and 6 by online MS/MS combining collisionally activated dissociation (CAD) and electron transfer dissociation (ETD). Offline MS/MS on Fourier-transform ion cyclotron resonance (FT-ICR) mass spectrometer with CAD and electron capture dissociation (ECD) was then utilized to achieve deep sequencing of mono-phosphorylated cTnT1 (35.2 kDa) with a high sequence coverage of 87%. Taken together, this study demonstrated the unique advantage of top-down MS in the comprehensive characterization of protein alternative splicing isoforms together with PTMs.
关键词	Cardiac troponin Heart disease Proteoform Top-down proteomics Collisionally activated dissociation Electron-transfer dissociation Electron-capture dissociation
DOI	10.1007/s13361-018-1925-y
收录类别	SCI
语种	英语
资助项目	NIH[R01 HL109810] ; NIH[HL096971] ; NIH[R01 GM117058] ; NIH[S10 OD018475]
WOS研究方向	Biochemistry & Molecular Biology ; Chemistry ; Spectroscopy
WOS类目	Biochemical Research Methods ; Chemistry, Analytical ; Chemistry, Physical ; Spectroscopy
WOS记录号	WOS:000435538300021
出版者	SPRINGER
引用统计	被引频次：13[WOS] [WOS记录] [WOS相关记录]
文献类型	期刊论文
条目标识符	http://119.78.100.204/handle/2XEOYT63/5108
专题	中国科学院计算技术研究所期刊论文_英文
通讯作者	Ge, Ying
作者单位	1.Univ Wisconsin, Dept Cell & Regenerat Biol, Madison, WI 53705 USA 2.Univ Wisconsin, Human Prote Program, Madison, WI 53705 USA 3.Shandong Prov Hosp, Dept Cardiol, Jinan 250021, Shandong, Peoples R China 4.Chinese Acad Sci, Inst Comp Technol, Beijing 100190, Peoples R China 5.Univ Wisconsin, Dept Chem, 1101 Univ Ave, Madison, WI 53706 USA 6.Univ Wisconsin, Dept Anim Sci, Madison, WI 53706 USA
推荐引用方式 GB/T 7714	Lin, Ziqing,Guo, Fang,Gregorich, Zachery R.,et al. Comprehensive Characterization of Swine Cardiac Troponin T Proteoforms by Top-Down Mass Spectrometry[J]. JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY,2018,29(6):1284-1294.
APA	Lin, Ziqing.,Guo, Fang.,Gregorich, Zachery R..,Sun, Ruixiang.,Zhang, Han.,...&Ge, Ying.(2018).Comprehensive Characterization of Swine Cardiac Troponin T Proteoforms by Top-Down Mass Spectrometry.JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY,29(6),1284-1294.
MLA	Lin, Ziqing,et al."Comprehensive Characterization of Swine Cardiac Troponin T Proteoforms by Top-Down Mass Spectrometry".JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY 29.6(2018):1284-1294.