Research And Application For Fluorescent Peptide Probes In The Detection Of Bioactive Molecule

Fluorescence technologies are powerful research tools for biochemical and cellular studies. With the aid of modern epifluorescence, confocal, and multiple photon microscopes, in combination with fluorescence probes, fluorescence-labeled molecules, and enzymes can be visualized and measured with precision and clarity. Recently, many novel peptide-based fluorescent molecular probes have been developed for in vivo biomedical imaging. To report specific information of biological targets, the probes were individually designed according to the unique property or functions of their targets. These peptide-based probes can be classified into targeting, crosslinking, and enzyme-activatable probes. Several of them have been tested in various in vitro and in vivo models, and the obtained imaging information has been applied to disease detection, medical diagnosis, and drug evaluations.Fluorescence resonance energy transfer (FRET) is one of the most powerful and widely used fluorescence techniques available for probing structure and dynamics in media. FRET is a powerful spectroscopic technique that allows biologically relevant distances between 20 and 80 ? to be quantified under physiological conditions with near-angstrom resolution due to the strong distance dependence of the transfer process. The efficiency of FRET is dependent upon donor-acceptor proximity and spectral overlap, whether the acceptor partner is fluorescent or not. The use of quenching acceptors is becoming increasingly popular in FRET systems. The principal advantage that these molecules offer over their fluorescent counterparts is the elimination of background fluorescence originating from direct acceptor excitation or re-emission. As a photoluminescent quencher, gold nanoparticles (AuNPs) can ultra-efficiently quench the molecular-excitation energy in chromophore-AuNPs composites. AuNPs have been of great interest because of their high extinction coefficient and a broad absorption spectrum in a visible light that is overlapped with the emission wavelength of usual energy donors. In comparison with the organic quencher, AuNPs have unique structural and optical properties for new applications in biosensing and molecular engineering.Enzyme has specific biological activity. It play important role in maintain the normal function of the body and other life activities. Matrix metalloproteinases (MMPs), also called matrixins, are a family of zinc-dependent neutral endopeptidases whose main function is degradation of the extracellular matrix. These enzymes are present in normal healthy individuals and have been shown to play an important role in many physiological processes. The main interest in the MMPs is due to their involvement in certain disease states in which the extracellular matrix breakdown is a key feature. Such diseases include rheumatoid arthritis, periodontal disease and cancer.Lots of enzyme-activatable probes are based on the FRET principle, however, probe for tow different kinds of proteases'simultaneous detection isn't reported. We design Au NPs as a quencher module of fluorescent probe for hydrolysis of amide bonds in peptides caused by MMP-2 (Matrix Metalloproteinase-2) and MMP-7(Matrix Metalloproteinase-7). In the present study, high sensitivity was obtained using the fluorescent chelated europium and 7-amino-4-methylcoumarin as label, internally quenched by suitable quencher Au NPs and released upon enzymatic reaction. This approach would allow sensitive monitoring of MMP-2 and MMP-7 activities in normal and cancerous cell cultures using FRET pair-labeled peptide substrate.Probes that can actively measure changes in the redox state of a given environment via fluorescence would circumvent limitations of current approaches such as the use of specialized equipment or, in the case of making in vivo measurements, irreversible cellular damage. Herein, we present the design and utility of a peptidic probe capable of accurately measuring environmental redox potential. When oxidized, the probe adopts a conformation in which 11-MUA-LAuND is in close spatial proximity to Dabcyl, limiting the sensitized emission process. When the probe is reduced, it adopts random coil conformation that spatially separates 11-MUA-LAuND and Dabcyl, allowing sensitized emission to occur. The fluorescence difference between the probe's oxidized and reduced states can be exploited to actively measure environmental redox potential.