基于电化学表面增强拉曼光谱的单细胞水平人DNA糖基化酶hOGG1活性检测研究

人8-羟基鸟嘌呤DNA糖基化酶（human 8-oxoguanine DNA glycosylase，hOGG1）能够特异性切除8-羟基鸟嘌呤修复氧化受损的DNA。细胞内hOGG1的表达异常及功能改变与包括恶性肿瘤在内的多种重大疾病的发生、发展及预后密切相关。但是，目前hOGG1活性检测方法灵敏度较低，且无法提供酶和DNA分子相互作用的实时动态信息。本项目拟结合电化学和表面增强拉曼光谱技术，利用hOGG1和受损DNA作用导致核酸分子电子传递性质改变的生物物理机制，借助SERS技术的单分子检测能力，发展高灵敏度、高选择性的原位分析新方法，实时研究hOGG1对受损DNA的碱基切除修复过程，以及在单细胞水平上考察hOGG1活性水平的异质性。本课题将为深入理解DNA修复过程的分子生物学机理，为相关癌症的早期预防和诊断提供新的更加有效的技术平台。

Human 8-oxoguanine (8-oxoG) DNA glycosylase (hOGG1) can repair the oxidative DNA damage by recognizing 8-oxoG:C base pairs, expulsing the 8-oxoG base and cleaving the DNA back bone. The abnormal expression of intracellular hOGG1 is associated with the genesis, progress and prognosis of many malignant diseases including cancer. However, existing methods for the detection of hOGG1 activity are limited by their low sensitivity and incapacity to provide the dynamic information between protein and DNA molecules in real-time. In this project, an ultrasensitive spectroelectrochemical strategy for hOGG1 activity assay is developed by coupling cyclic voltammetry and surface-enhanced Raman spectroscopy (SERS). The protocol relies on the inhibition of DNA chemistry after damage repair by hOGG1 proteins. The single-molecule sensitivity of SERS allows us to profile the heterogeneity of hOGG1 activity at the single cell level. This project will provide a novel and more effective platform for getting a deep insight into the molecular biological mechanism of DNA damage repair, and prevention and early diagnosis of the associated cancers.