Thermal effect on aequorea green fluorescent protein anionic and neutral chromophore forms fluorescence

© Springer Science+Business Media, LLC 2011.The emission behaviour of Aequorea green fluorescent protein (A-GFP) chromophore, in both neutral (N) and anionic (A) form, was studied in the temperature range from 20 °C to 75 °C and at pH=7. Excitation wavelengths of 399 nm and 476 nm were applied to probe the N and A forms environment, respectively. Both forms exhibit distinct fluorescence patterns at high temperature values. The emission quenching rate, following a temperature increase, is higher for the chromophore N form as a result of the hydrogen bond network weakening. The chromophore anionic form emission maximum is red shifted, upon temperature increase, due to a charge transfer process occurring after A form excitation.The author acknowledges financial support from Fundação para a Ciência e a Tecnologia through grant SFRH/BPD/26821/2006. Dr. V. Subramanian (University of Twente,NL) for providing the A-GFP gene, and Dr. F. Mergulhão (University of Porto, PT) for the support given on A-GFP expression and purification