title: Time-resolved luminescence biosensor for continuous activity detection of protein acetylation-related enzymes based on DNA-sensitized terbium(III) probes.
abstract: Protein acetylation of histone is an essential post-translational modification (PTM) mechanism in epigenetic gene regulation, and its status is reversibly controlled by histone acetyltransferases (HATs) and histone deacetylases (HDACs). Herein, we have developed a sensitive and label-free time-resolved luminescence (TRL) biosensor for continuous detection of enzymatic activity of HATs and HDACs, respectively, based on acetylation-mediated peptide/DNA interaction and Tb(3+)/DNA luminescent probes. Using guanine (G)-rich DNA-sensitized Tb(3+) luminescence as the output signal, the polycationic substrate peptides interact with DNA with high affinity and subsequently replace Tb(3+), eliminating the luminescent signal. HAT-catalyzed acetylation remarkably reduces the positive charge of the peptides and diminishes the peptide/DNA interaction, resulting in the signal on detection via recovery of DNA-sensitized Tb(3+) luminescence. With this TRL sensor, HAT (p300) can be sensitively detected with a wide linear range from 0.2 to 100 nM and a low detection limit of 0.05 nM. The proposed sensor was further used to continuously monitor the HAT activity in real time. Additionally, the TRL biosensor was successfully applied to evaluating HAT inhibition by two specific inhibitors, anacardic acid and C464, and satisfactory Z'-factors above 0.73 were obtained. Moreover, this sensor is feasible to continuously monitor the HDAC (Sirt1)-catalyzed deacetylation with a linear range from 0.5 to 500 nM and a detection limit of 0.5 nM. The proposed sensor is a convenient, sensitive, and mix-and-read assay, presenting a promising platform for protein acetylation-targeted epigenetic research and drug discovery.
authors: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University , Changsha, 410082, People's Republic of China.
authors: Yitao
authors: Y
authors: Han
authors: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University , Changsha, 410082, People's Republic of China.
authors: Hao
authors: H
authors: Li
authors: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University , Changsha, 410082, People's Republic of China.
authors: Yufang
authors: Y
authors: Hu
authors: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University , Changsha, 410082, People's Republic of China.
authors: Pei
authors: P
authors: Li
authors: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University , Changsha, 410082, People's Republic of China.
authors: Huixia
authors: H
authors: Wang
authors: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University , Changsha, 410082, People's Republic of China.
authors: Zhou
authors: Z
authors: Nie
authors: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University , Changsha, 410082, People's Republic of China.
authors: Shouzhuo
authors: S
authors: Yao
mesh_terms: D000107:Acetylation; D015374:Biosensing Techniques / Q000379:methods*; D004247:DNA / Q000737:chemistry*; D004277:DNA, Single-Stranded / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006147:Guanine / Q000737:chemistry; D051548:Histone Acetyltransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006655:Histone Deacetylases / Q000378:metabolism*; D006657:Histones / Q000378:metabolism*; D057230:Limit of Detection; D049408:Luminescent Agents / Q000737:chemistry*; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D010455:Peptides / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D013725:Terbium / Q000737:chemistry*; D013997:Time Factors
chemical_list: D004277:DNA, Single-Stranded; D004791:Enzyme Inhibitors; D006657:Histones; D049408:Luminescent Agents; D010455:Peptides; D013725:Terbium; D006147:Guanine; D004247:DNA; D051548:Histone Acetyltransferases; D006655:Histone Deacetylases