Monoclonal Anti-AMP Antibodies Are Sensitive and Valuable Tools for Detecting Patterns of AMPylation.
Monoclonal Anti-AMP Antibodies Are Sensitive and Valuable Tools for Detecting Patterns of AMPylation.
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DOI：
10.1016/j.isci.2020.101800
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发表时间：
2020-12-18
期刊：
影响因子：
5.8
通讯作者：
Itzen A
中科院分区：
文献类型：
--
作者：
Höpfner D;Fauser J;Kaspers MS;Pett C;Hedberg C;Itzen A
AMP化是一种翻译后修饰，其用腺苷一磷酸（AMP）修饰氨基酸侧链。最近，AMPylation作为感染和细胞稳态中的通用调节机制的作用已经出现，推动了对研究这种修饰的通用工具的需求。在这里，我们描述了三个单克隆抗AMP抗体（单克隆抗体）从小鼠，能够蛋白质骨架独立的识别AMP化，在变性（蛋白质印迹）以及本地（ELISA，IP）的应用，从而优于以前报道的工具。这些抗体对AMP修饰具有高度敏感性和特异性，突出了它们作为新靶标鉴定以及已知靶标验证工具的潜力。有趣的是，将抗AMP mAb应用于各种癌细胞系揭示了以前未描述的广泛和多样的AMP化模式。总之，这些抗AMP单克隆抗体将进一步推进目前对AMP化和修饰靶点谱的理解。通过合成AMP化肽生成鼠单克隆抗AMP抗体应用中的表征蛋白质印迹、ELISA和免疫沉淀对AMP化的灵敏和特异性识别不依赖于蛋白质骨架扩增用于检测和富集AMP化的工具箱生物化学;生物分子;分子生物学;生物技术
AMPylation is a post-translational modification that modifies amino acid side chains with adenosine monophosphate (AMP). Recently, a role of AMPylation as a universal regulatory mechanism in infection and cellular homeostasis has emerged, driving the demand for universal tools to study this modification. Here, we describe three monoclonal anti-AMP antibodies (mAbs) from mouse that are capable of protein backbone-independent recognition of AMPylation, in denatured (western blot) as well as native (ELISA, IP) applications, thereby outperforming previously reported tools. These antibodies are highly sensitive and specific for AMP modifications, highlighting their potential as tools for new target identification, as well as for validation of known targets. Interestingly, applying the anti-AMP mAbs to various cancer cell lines reveals a previously undescribed broad and diverse AMPylation pattern. In conclusion, these anti-AMP mABs will further advance the current understanding of AMPylation and the spectrum of modified targets. Generation of murine monoclonal anti-AMP antibodies via synthetic AMPylated peptide Characterization in the applications western blot, ELISA, and immunoprecipitation Sensitive and specific recognition of AMPylation independent of protein backbone Expansion of toolbox for the detection and enrichment of AMPylation Biochemistry; Biomolecules; Molecular Biology; Biotechnology
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影响因子：
2.9
作者：
Gibson, Bryan A.;Conrad, Lesley B.;Kraus, W. Lee
通讯作者：
Kraus, W. Lee
DOI：
10.1111/j.1432-1033.1978.tb12001.x
发表时间：
1978-01-01
期刊：
EUROPEAN JOURNAL OF BIOCHEMISTRY
影响因子：
--
作者：
BREDEHORST, R;FERRO, AM;HILZ, H
通讯作者：
HILZ, H
影响因子：
15
作者：
Grammel, Markus;Phi Luong;Orth, Kim;Hang, Howard C.
通讯作者：
Hang, Howard C.
DOI：
10.1074/jbc.m111.227603
发表时间：
2011-09-16
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Mattoo S;Durrant E;Chen MJ;Xiao J;Lazar CS;Manning G;Dixon JE;Worby CA
通讯作者：
Worby CA
影响因子：
4.6
作者：
Khater, Shradha;Mohanty, Debasisa
通讯作者：
Mohanty, Debasisa