Ultrafast proton transfer dynamics on a short H-bond in a protein were studied through the time-resolved fluorescence of the S65T/H148D green fluorescent protein (GFP) mutant. In response to the change in chromophore pK(a) upon excitation, the donor-proton-acceptor structure evolves on a sub- 100 fs time scale, followed by picosecond time scale vibrational cooling and host structure reorganization.