Study On The Activity Of Protein Kinase By Photoelectrochemical

Protein phosphorylation by kinases plays important regulatory roles in diverse cellular functions including the regulation of metabolism, gene transcription, cell cycle progression, apoptosis and signal transduction. Thus, simple and sensitive assays for determining protein kinase activity are urgently needed. Photoelectrochemical approach is a newly developed detection method which contains the merits of reduced background signal and high sensitization. Selectively recognition of the phosphorylated peptide and signal amplification are two important factors for sensitive measurement of protein kinase activity and their potential inhibitors screening. Herein, three novel PEC biosensors were fabricated for sensitive and accurate protein kinase activety analysis. They are not only helpful for the clinical diagnosis and targeted therapy, but also necessary for further understanding the molecular mechanism of signal transduction.Based on localized surface plasmon resonance enhancement and dye sensitization, a novel visible- light photoelectrochemical biosensor was fabricated for highly sensitive analysis of protein kinase activity with ultralow background. The adsorbed [Ru(bpy)3]2+can harvest visible light to produce excited electrons that inject into the Ti O2 conduction band to form photocurrent under visible light irradiation. In addition, the photocurrent efficiency was further improved by the localized surface plasmon resonance of Gold Nanoparticles(Au NPs) under the irradiation of visible light. Moreover, because of the excellent conductivity and large surface area of Au NPs, the photocurrent was significantly amplified, affording an extremely sensitive PEC analysis of kinase activity with ultralow background signals. The detection limit of as- proposed PEC biosensor was0.005 U m L-1( S/N = 3). The biosensor also showed excellent performances for quantitative kinase inhibitor screening.A facial all- in- one multifunctional MOFs as both transducer and amplifier in photoelectrochemical biosensor was fabricated for highly sensitive protein kinase activity analysis and inhibitor screening. It was first proposed that [Ru(bpy)3]2+ dye loaded Ui O-66 probes were assembled on the modified electrode through the high affinity between Zr- O clusters in Ui O- 66 and phosphate groups in phosphorylated kemptide. The detection limit of as- proposed photoelectrochemical biosensor was 0.0049 U m L-1( S/N = 3). The biosensor can also be applied for quantitative kinase inhibitor screening and PKA activities detection in MCF- 7 cell lysates.Based on exciton- plasmon interactions between CdS quantum dots and Au NPs, a novel photoelectrochemical biosensor was proposed for protein kinase activity analysis. Due to their natural absorption overlap, the exciton of the QDs and the plasmon of Au NPs could be stimulated of exciton- plasmon interactions, and the photocurrent was used for PKA activity evaluation. The biosensor can also be successfully applied for PKA activities detection in MCF- 7 cell lysates.