Renilla reniformis bioluminescence: luciferase-catalyzed production of nonradiating excited states from luciferin analogues and elucidation of the excited state species involved in energy transfer to Renilla green fluorescent protein
A number of coelenterate-type luciferin analogues with structural changes in the p-hydroxyphenyl and p-hydroxybenzyl substituents have been synthesized. During chemiluminescence, each of the analogues produces a blue emission arising from the singlet excited state of the corresponding oxyluciferin monoanion. During bioluminescence two emissions are observed with coelenterate-type luciferin and some of its analogues. One of these arises fom the amide monoanion (lambda/sub max/ approximately 480 nm) and the other arises from the neutral species of oxyluciferin (lambda/sub max/ approximately 395 nm). Certain analogues produce both emissions, while others produce only the near-ultraviolet emission. Structural changes in the p-hydroxyphenyl substituent result in complete or nearly complete elimination of emission from the monoanion, resulting in over a 100-fold reduction in bioluminescence quantum yield. Structural changes in the p-hydroxybenzyl substituent do not have a significant effect on the emission spectrum but decrease the luciferase turnover number approximately 25-fold. The large decrease in the bioluminescence quantum yield observed with some of the analogues can be overcome by addition of green fluoroescent protein (GFP). GFP forms a rapid equilibrium complex with luciferase and is known to function in this system as an efficient energy-transfer acceptor (Ward, W. W., and Cormier, M. J. (1978) Phtochem. Photobiol. 27, 389-396). Spectral analyses have shown that radiationless energy transfer occurs from the singlet excited state of the oxyluciferin monoanion and not from the neutral excited species. The energy-transfer data suggests that the luciferase-bound monoanion singlet excited state can be quenched by solvent and/or protein functional groups. Energy transfer to GFP can apparently favorably compete with this quenching process. Lifetime measurements have shown that the rate of energy transfer must be at least 3 x 10/sup 8/ s/sup -1/.
- Research Organization:
- Univ. of Georgia, Athens
- OSTI ID:
- 5324594
- Journal Information:
- Biochemistry; (United States), Vol. 18:11
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
CNIDARIA
LUMINESCENCE
LUCIFERASE
ENZYME ACTIVITY
LUCIFERIN
EXCITED STATES
ENERGY TRANSFER
EMISSION SPECTRA
EXPERIMENTAL DATA
FLOWSHEETS
FLUORESCENCE
GRAPHS
PROTEINS
TABLES
ALBUMINS
DATA
DATA FORMS
DIAGRAMS
ENERGY LEVELS
ENZYMES
INFORMATION
NUMERICAL DATA
ORGANIC COMPOUNDS
OXIDASES
OXIDOREDUCTASES
SPECTRA
550200* - Biochemistry