title: pH-sensitive pHluorins as a molecular sensor for in situ monitoring of enzyme-catalyzed prodrug activation.
abstract: This work examines the feasibility of using a pH-sensitive fluorescent protein as a molecular reporter for enzyme-catalyzed prodrug activation reaction. Specifically, a ratiometric pHluorins was examined for detection of the activity of horseradish peroxidase (HRP) for the activation of indole-3-acetic acid. The pHluorins and HRP were conjugated chemically, forming a biocatalyst with a self-reporting function. Results showed that the characteristic fluorescence intensity ratio of the conjugate shifted from 1.47 to 1.40 corresponding to the progress of the prodrug activation reaction. The effectiveness of applying the conjugate for inhibition of the growth of Bcap-37 cells was also demonstrated simultaneously with reaction monitoring. The results reveal a very promising approach to realizing in situ monitoring of enzyme activities based on pH shifting for enzyme-based prodrug therapy applications.
authors.affiliation: Biomedical Nanotechnology Center, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China.
authors.forename: Hui
authors.initials: H
authors.lastname: Liu
authors.affiliation: Biomedical Nanotechnology Center, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China.
authors.forename: Xiaodan
authors.initials: X
authors.lastname: Cao
authors.affiliation: Biomedical Nanotechnology Center, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China.
authors.forename: Ping
authors.initials: P
authors.lastname: Wang
authors.affiliation: Biomedical Nanotechnology Center, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China.
authors.forename: Xingyuan
authors.initials: X
authors.lastname: Ma
mesh_terms: D055162:Biocatalysis; D015374:Biosensing Techniques; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004789:Enzyme Activation; D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D006735:Horseradish Peroxidase / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007210:Indoleacetic Acids / Q000737:chemistry / Q000378:metabolism*; D015335:Molecular Probes / Q000737:chemistry* / Q000378:metabolism; D011355:Prodrugs / Q000737:chemistry / Q000378:metabolism*
chemical_list: D007210:Indoleacetic Acids; D015335:Molecular Probes; C485187:PHluorin; D011355:Prodrugs; D049452:Green Fluorescent Proteins; C030737:indoleacetic acid; D006735:Horseradish Peroxidase
keywords: Bcap-37; HRP; IAA; enzyme/prodrug therapy; ratiometric pHluorins