roGFP

roGFP stands for redox-sensitive Green Fluorescent Protein. It is a genetically encoded biosensor used to monitor the redox state of specific cellular compartments or the entire cell. roGFP is a fusion protein consisting of two copies of GFP linked by a peptide linker. Crucially, between these two GFP molecules is a single cysteine residue that is susceptible to oxidation and reduction. This redox-sensitive cysteine residue alters the fluorescence properties of the protein. In a reduced state, roGFP exhibits a specific fluorescence emission spectrum, while in an oxidized state, this spectrum shifts. This change in fluorescence allows researchers to detect and quantify redox changes.

The ratiometric nature of roGFP is key to its utility. By measuring fluorescence at two different excitation wavelengths (typically around 400 nm and 490 nm), a ratio of fluorescence intensities can be calculated. This ratio directly correlates with the redox state of the environment where roGFP is localized. For example, a higher ratio might indicate a more reduced environment, while a lower ratio suggests a more oxidized environment.

Researchers can target roGFP to specific organelles, such as mitochondria, the endoplasmic reticulum, or the cytoplasm, by fusing it to organelle-specific localization signals. This allows for the investigation of redox dynamics in different cellular compartments. roGFP has been widely used to study various cellular processes, including oxidative stress, apoptosis, and metabolic pathways, providing valuable insights into the role of redox balance in cellular function and disease.