Spectroscopy and microscopy of the autofluorescent protein DsRed from Discosoma genus coral

Mircea Cotlet; Satoshi Habuchi; Frans Carl De Schryver; Johan Hofkens

doi:10.1117/12.531313

14 June 2004 Spectroscopy and microscopy of the autofluorescent protein DsRed from Discosoma genus coral

Mircea Cotlet, Satoshi Habuchi, Frans Carl De Schryver, Johan Hofkens

Proceedings Volume 5329, Genetically Engineered and Optical Probes for Biomedical Applications II; (2004) https://doi.org/10.1117/12.531313
Event: Biomedical Optics 2004, 2004, San Jose, CA, United States

Abstract

We report on the fluorescence dynamics of a red fluorescent protein DsRed from the coral Discosoma genus by means of ensemble and single molecule fluorescence spectroscopy. Single molecule experiments performed on 543-nm excitation point to the existence of DsRed as a tetramer and reveal the presence of a no/off blinking phenomenon in the millisecond time range. Collective effects involving the red chromophores within the individual tetramers were observed. Time-resolved fluorescence data reveal the presence of a population of 25 % of the immature green chromophores which relates to tetramers containing only this immature green form and which is responsible for the weak fluorescence emitted by DsRed at 500-nm when excited at 460-nm. The remaining 75 % of the immature green chromophores are involved in a FRET process to the red chromophores within the tetramers that contain them. Using time-resolved detection and spectroscopy at single molecule level we were able to demonstrate the presence of a photoconversion process of the red chromophore emitting at 583-nm into a super red species that emits weakly at 595-nm. The same phenomenon is further corroborated at the ensemble level with the observation of the creation of a super red form and a blue absorbing species upon irradiation with 532-nm pulsed light at high excitation power.

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Mircea Cotlet, Satoshi Habuchi, Frans Carl De Schryver, and Johan Hofkens "Spectroscopy and microscopy of the autofluorescent protein DsRed from Discosoma genus coral", Proc. SPIE 5329, Genetically Engineered and Optical Probes for Biomedical Applications II, (14 June 2004); https://doi.org/10.1117/12.531313

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KEYWORDS

Chromophores

Luminescence

Molecules

Proteins

Fluorescence spectroscopy

Absorption

Green fluorescent protein

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