相关实验视频

Updated: Feb 23, 2026

Deacetylation Assays to Unravel the Interplay between Sirtuins SIRT2 and Specific Protein-substrates

Deacetylation Assays to Unravel the Interplay between Sirtuins SIRT2 and Specific Protein-substrates

Published on: February 27, 2016

基因编码的光探测器用于检测活细胞中的Sirtuins

Weimin Xuan¹, Anzhi Yao¹, Peter G Schultz¹

¹Department of Chemistry and Skaggs Institute for Chemical Biology, the Scripps Research Institute , 10550 N Torrey Pines Road, La Jolla, California 92037, United States.

Journal of the American Chemical Society

|September 1, 2017

概括

研究人员开发了一种新的光探测器, 这种工具可以监测对各种生物过程至关重要的Sirtuin酶.

科学领域:

生物化学
分子生物学
细胞生物学

背景情况:

赛尔图因是NAD+依赖的蛋白质脱乙酶,调节各种生物功能.
对解读细胞过程和疾病机制至关重要.

研究的目的:

开发一种新型的基因编码光探测器,用于实时监测活细胞中的Sirtuin活性.
验证探针对各种人类素异型的反应及其适用于不同翻译后修改.

主要方法:

设计和合成一种基因编码的光探针 (EGFP-K85AcK),利用EGFP中的氨酸残留物.
在表达人类素的大肠杆菌 ΔcobB 和 HEK293T 细胞中测试探针活性.
使用探针的mCherry融合来监测SIRT1和SIRT2活动.

主要成果:

EGFP-K85AcK探针成功对多种人体素 (SIRT1,SIRT2,SIRT3,SIRT5) 进行了反应.
在HEK293T细胞中实现了SIRT1和SIRT2活性的直接监测.
这种方法扩展到其他光蛋白,并证明了在额外的氨酸修饰物上对氨酸的活性.

结论:

一个新的光探测器可以在体内敏感和特定地检测素活性.
这种工具有助于研究素的酶活性及其在细胞调节中的作用.
该探测平台可适应各种与素相关的研究和药物发现工作.

更多相关视频

Single-Cell Quantification of Protein Degradation Rates by Time-Lapse Fluorescence Microscopy in Adherent Cell Culture

Single-Cell Quantification of Protein Degradation Rates by Time-Lapse Fluorescence Microscopy in Adherent Cell Culture

Published on: February 4, 2018

Monitoring of Ubiquitin-proteasome Activity in Living Cells Using a Degron dgn-destabilized Green Fluorescent Protein GFP-based Reporter Protein

Monitoring of Ubiquitin-proteasome Activity in Living Cells Using a Degron dgn-destabilized Green Fluorescent Protein GFP-based Reporter Protein

Published on: November 10, 2012

相关概念视频

Labeling DNA Probes

Labeling DNA Probes

DNA probes are fragments of DNA labeled with a reporter tag to enable their detection or purification. The resulting labeled DNA probes can then hybridize to target nucleic acid sequences through complementary base-pairing, and may be used to recover or identify these regions.
Radioisotopes, fluorophores, or small molecule binding partners like biotin or digoxigenin, are the most widely used reporter tags for labeling DNA probes. These labels can be attached to the probe DNA molecule via...

Reporter Genes

Reporter Genes

Reporter genes are a type of protein-coding gene that are often tagged to a gene of interest. Once inside a target cell, reporter genes usually produce visually identifiable characteristics like fluorescence and luminescence when expressed along with the gene of interest. Thus, reporter genes “report” the presence or absence of genes of interest in an organism, determine the gene expression pattern, or track the physical location of a DNA segment or protein in the cell.

Protein Dynamics in Living Cells

Protein Dynamics in Living Cells

Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...