Quenching of tryptophan fluorescence of firefly luciferase by substrates.
The interaction of firefly luciferase with substrates (luciferin and MgATP) by steady-state and time-resolved fluorescence is studied. The efficient quenching of tryptophan fluorescence of the active enzyme takes place upon its binding with substrates. In the presence of ATP the quenching is of dynamic type and is caused by structural changes in the protein molecule upon ATP binding. A model is proposed in which the complex has smaller fluorescence quantum yield than the free enzyme because of partial quenching of tryptophan fluorescence by the new microenvironment. Quenching of tryptophan fluorescence by luciferin due to the efficient energy transfer from tryptophan to luciferin is discussed. The calculated distance between Trp-419 and luciferin for the L. mingrelica luciferase in the enzyme-substrate complex is less than 12 A.
Cherednikova EYu
Physics Department and International Laser Center, Moscow, Russia.
Chikishev AYu
Dementieva
E I
EI
Kossobokova
O V
OV
Ugarova
N N
NN
D000255:Adenosine Triphosphate / Q000737:chemistry*; D000818:Animals; D001517:Coleoptera; D049454:Firefly Luciferin / Q000737:chemistry*; D005453:Fluorescence; D008156:Luciferases / Q000737:chemistry*; D013050:Spectrometry, Fluorescence / Q000379:methods; D013379:Substrate Specificity; D014364:Tryptophan / Q000737:chemistry
D049454:Firefly Luciferin; D014364:Tryptophan; D000255:Adenosine Triphosphate; D008156:Luciferases