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12th International Symposium on Bioluminescence & Chemiluminescence |
Symposium abstracts:
Tisi, Laurence C., Law, Gim Hoong, Gandelmann, Olga
A., Lowe, Christopher R., Murray, James A.H.
Institute of Biotechnology, Tennis Court Road, Cambridge CB2 1QT, UK
Email: lct11@mole.bio.cam.ac.uk
The characteristic yellow-green light emission catalysed by the luciferase from the firefly Photinus pyralis at pH 7.8 is red-shifted as the pH of the reaction is lowered. The basis of this "bathochromic shift" is thought to be the protonation of the emitter oxyluciferin. However, related beetle luciferases do not have a bathochromic shift (including green emitting luciferases). Thus it appears that depending on the luciferase, the ease with which the emitter can be protonated is dramatically different. This in principle, could be explained by whether or not a particular luciferase has an appropriately orientated base in juxtaposition to the emitter. However, presented here is evidence there may be an alternative mechanism responsible.
We have identified several amino acid substitutions in the Photinus pyralis luciferase that greatly reduce the bathochromic shift. Many of the substitutions do not remove or add a base; in fact some substitutions are of hydrophobic residues for other hydrophobic residues. A common feature of these substitutions however, is that they all increase the thermostability of the enzyme. Thus there appears to be a link between the bathochromic shift and luciferase thermostability. This suggests that a conformational change may be the basis of the bathochromic shift.
This
is a preprint of an article accepted for publication in Luminescence: Copyright
2001 John
Wiley & Sons, Ltd (Wiley website)