Use of the regulatory subunit of the cAMP dependent protein kinase (PKA) from dictyostelium for cAMP measurements

摘要

The invention relates to the use of the regulatory subunit (R) of the CAMP dependent protein kinase (PKA) from Dictyostelium discoideum for cAMP detection. It includes constructs for expression of the R-subunit in E. coli and fusion to green fluorescent proteins (GFP). Fluorescence energy transfer is used as a way to monitor cAMP binding, either by using fluorescently labelled cAMP or cGMP, or by using mutant GFPs with modified absorption and emission spectra. FRET changes upon cAMP binding will allow measurement of cAMP level either in vitro or within living cells.