Bifunctional role of leucine 300 of firefly luciferase in structural rigidity.Firefly luciferase is susceptible to thermal inactivation, thereby its intracellular half-life decreased.  Previous reports indicated that L300R mutation (LRR mutant) in E354R/Arg356 double mutant (ERR mutant) from Lampyris turkestanicus luciferase has increased its thermal stability and rigidity through induction of some ionic bonds with Asp 270 and 271.  In this study, we investigated the effects of this residue to find the truth behind an extraordinary increase in thermal stability and rigidity of luciferase after replacement of leucine 300 by arginine based on previous reports.  In spite of increase of intrinsic fluorescence of the mutants a slight increase in thermostability and retention of kinetic properties was observed.  Based on our results, we can conclude that L300R mutation in LRR mutant accompanying with alteration in a flexible loop (352-359) increased thermostability and rigidity of luciferase.
THERM