Xia A, Jiang S, Zhu J, Shen Y, Lu Z, Huang W
National Laboratory of Molecular and Biomolecular Electronics, Southeastern University, 210096 Nanjing.
Guang Pu Xue Yu Guang Pu Fen Xi. 2001 Aug;21(4):435-7.
The photoconversion process of recombinant green fluorescent protein (rGFP) was investigated by two-photon excitation. The results indicated that the rGFP had very strong two-photon excitation fluorescence. The changes of two-photon-induced fluorescence polarization suggest that there is a proton transfer process between two different proton states of rGFP chromophores. The conformation of rGFP chromophores could be changed upon illumination, which partly block the energy transfer processes from amino acid residues to chromophores in rGFP, and result in the decrease of two-photon-induced fluorescence intensity. The fluorescence from amino acid residues in rGFP was also observed by three-photon excitation, which resulted from the blocked amino acid residues in rGFP. These results suggested that it is necessary to optimize rGFP excitation and detection for quantitative fluorescence microscopy.
通过双光子激发研究了重组绿色荧光蛋白（rGFP）的光转换过程。结果表明，rGFP具有非常强的双光子激发荧光。双光子诱导荧光偏振的变化表明，rGFP发色团的两种不同质子状态之间存在质子转移过程。光照后rGFP发色团的构象可能会发生变化，这部分地阻断了rGFP中从氨基酸残基到发色团的能量转移过程，并导致双光子诱导荧光强度降低。通过三光子激发也观察到了rGFP中氨基酸残基发出的荧光，这是由rGFP中被阻断的氨基酸残基引起的。这些结果表明，有必要优化rGFP的激发和检测以用于定量荧光显微镜检查。
