Click-Type Protein-DNA Conjugation for Mn2+ Imaging in Living Cells.,Analytical Chemistry

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Click-Type Protein-DNA Conjugation for Mn2+ Imaging in Living Cells.
Analytical Chemistry ( IF 6.7 ) Pub Date : 2019-07-04 00:00:00 , DOI: 10.1021/acs.analchem.9b02191
Shanfang Hu ₁ , Jinghui Zhang ₁ , Rui Tang ₁ , Jiahui Fan ₁ , Huiqiong Liu ₁ , Wenyuan Kang ₁ , Chunyang Lei ₁ , Zhou Nie ₁ , Yan Huang ₁ , Shouzhuo Yao ₁

Affiliation

A click-type protein–DNA conjugation, named as MnDDC (Mn²⁺-activated DCV-DNA conjunction), is presented, where DCV (rep protein of duck circovirus) and its target DNA work as the modular blocks to rapidly and effectively generate Mn²⁺-dependent and site-specific protein–DNA linkage. On the basis of MnDCC, a fluorescent Mn²⁺ biosensor composed of DCV and a molecular beacon, was developed for rapid sensing of Mn²⁺ within 2 min with nanomolar sensitivity. Using the proposed biosensor, not only analysis of Mn²⁺ in real samples (e.g., serum and food), but also wash-free fluorescent imaging of Mn²⁺ in extracellular environment and cytoplasm have been achieved. Moreover, the MnDDC-based sensor was proved to be a powerful tool for visualization of Mn²⁺ during exploration of the associated cytotoxicity in living neural cells, which is helpful to reveal the cellular responses toward the disordered homeostasis of Mn²⁺ in both extracellular and intracellular microenvironments.

中文翻译：

用于活细胞中Mn2 +成像的点击型蛋白质-DNA偶联。

提出了一种单击型蛋白质与DNA的结合，称为MnDDC（Mn ²⁺激活的DCV-DNA结合体），其中DCV（鸭圆环病毒的rep蛋白）及其靶DNA作为模块块起作用，可以快速有效地产生Mn ²⁺依赖性和位点特异性蛋白质-DNA连锁。基于MnDCC，开发了由DCV和分子信标组成的荧光Mn ²⁺生物传感器，用于以纳摩尔灵敏度在2分钟内快速检测Mn ²⁺。使用所提出的生物传感器中，不仅锰的分析²⁺实际样品（例如，血清和食物）中，而且还洗游离锰的荧光成像²⁺已经在细胞外环境和细胞质中获得了成功。此外，基于MnDDC的传感器被证明是探索活神经细胞相关细胞毒性过程中Mn ²⁺可视化的有力工具，这有助于揭示细胞对Mn2 ⁺在两个细胞外稳态中无序动态的反应。和细胞内的微环境。

更新日期：2019-07-04

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