RECOMBINANT FLUORESCENT SENSOR OF HYDROGEN PEROXIDE HyPer FUSED WITH ADAPTOR PROTEIN Ruk/CIN85: DESIGNING OF EXPRESSION VECTOR AND ITS FUNCTIONAL CHARACTERIZATION

The aim of this study was to design the expression vector encoding fluorescent sensor of hydrogen peroxide HyPer fused with adaptor protein Ruk/CIN85 as well as to check its subcellular distribution and ability to sense hydrogen peroxide. It was demonstrated that in transiently transfected HEK293 and MCF-7 cells Ruk/CIN85-HyPer is concentrated in dot-like vesicular structures of different size while HyPer is diffusely distributed throughout the cell. Using live cell fluorescence microscopy we observed gradual increase in hydrogen peroxide concentration in representative vesicular structures during the time of experiment. Thus, the developed genetic construction encoding the chimeric Ruk/CIN85-HyPer fluorescent protein represents a new tool to study localized H2O2 production in living cells.