Enzymatic immuno-assembly of gold nanoparticles for visualized activity screening of histone-modifying enzymes.
Enzymatic immuno-assembly of gold nanoparticles for visualized activity screening of histone-modifying enzymes.
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DOI：
10.1021/ac203385v
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发表时间：
2012-04
影响因子：
7.4
通讯作者：
Zhen Zhen-Zhen;Li-Juan Tang;Hao Long;Jian-hui Jiang
中科院分区：
文献类型：
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作者：
Zhen Zhen-Zhen;Li-Juan Tang;Hao Long;Jian-hui Jiang
组蛋白修饰酶的活性筛选对于表观遗传学研究以及临床诊断和治疗至关重要。一种新的生物传感策略已经发展为敏感和选择性检测组蛋白修饰酶及其抑制剂。这种策略依赖于抗体介导的组装金纳米颗粒（AuNPs）装饰与底物肽，进行酶修饰的组蛋白修饰酶。这种设计允许使用没有任何标记的抗体进行酶活性的目视和均相测定，这避免了对抗体进行预官能化的要求，并提供了改进的测定简单性和通量。此外，与基于酶偶联测定的现有技术相比，使用基于抗体的修饰肽识别可以提供改进的特异性。我们已经证明了这种策略，使用组蛋白甲基转移酶作用于组蛋白H3（赖氨酸4）和组蛋白乙酰转移酶作用于组蛋白H3（赖氨酸14）。结果表明，金纳米颗粒的吸收峰特征随着酶活性的增加而动态降低，伴随着可见的颜色衰减，并且两种酶都很容易达到亚纳摩尔的检测限。该方法为高灵敏度、高选择性的酶活性及其抑制剂的筛选提供了一个方便、可靠的可视化平台。
Activity screening of histone-modifying enzymes is of paramount importance for epigenetic research as well as clinical diagnostics and therapeutics. A novel biosensing strategy has been developed for sensitive and selective detection of histone-modifying enzymes as well as their inhibitors. This strategy relies on the antibody-mediated assembly of gold nanoparticles (AuNPs) decorated with substrate peptides that are subjected to enzymatic modifications by the histone-modifying enzymes. This design allows a visual and homogeneous assay of the enzyme activity using antibodies without any labels, which circumvents the requirements to prefunctionalize the antibody and affords improved assay simplicity and throughput. Additionally, the use of antibody-based recognition of modified peptides could offer improved specificity as compared with existing techniques based on the enzyme coupled assay. We have demonstrated this strategy using a histone methyltransferase acting on histone H3 (Lys 4) and a histone acetyltransferase acting on histone H3 (Lys 14). The results reveal that the absorption peak characteristic for AuNPs decreases dynamically with increasing activity of the enzymes with concomitant visualizable color attenuation, and subnanomolar detection limits are readily achieved for both enzymes. The developed strategy can thus offer a robust and convenient visualized platform for screening the enzyme activities and their inhibitors with high sensitivity and selectivity.