The structural origin and biological function of pH-sensitivity in firefly luciferases

Abstract

Firefly luciferases are called pH-sensitive because their bioluminescence spectra display a typical red-shift at acidic pH, higher temperatures, and in the presence of heavy metal cations, whereas other beetle luciferases (click beetles and railroadworms) do not, and for this reason they are called pH-insensitive. Despite many studies on firefly luciferases, the origin of pH-sensitivity is far from being understood. This subject is revised in view of recent results. Some substitutions of amino-acid residues influencing pH-sensitivity in firefly luciferases have been identified. Sequence comparison, site-directed mutagenesis and modeling studies have shown a set of residues differing between pH-sensitive and pH-insensitive luciferases which affect bioluminescence colors. Some substitutions dramatically affecting bioluminescence colors in both groups of luciferases are clustered in the loop between residues 223-235 (Photinus pyralis sequence). A network of hydrogen bonds and salt bridges involving the residues N229-S284-E311-R337 was found to be important for affecting bioluminescence colors. It is suggested that these structural elements may affect the benzothiazolyl side of the luciferin-binding site affecting bioluminescence colors. Experimental evidence suggest that the residual red light emission in pH-sensitive luciferases could be a vestige that may have biological importance in some firefly species. Furthermore, the potential utility of pH-sensitivity for intracellular biosensing applications is considered.

A Time‐Dependent Density Functional Theory Investigation on the Origin of Red Chemiluminescence

“…[45] A network of hydrogen bonds formed by amino residues surrounding the active site and emitter is controlled by pH-sensitive or pH-insensitive luciferases, generating open or closed conformations. The open or close conformation is related to two important amino-acid residues in the active site.…”

Study on the Effects of Intermolecular Interactions on Firefly Multicolor Bioluminescence

“…Also, control of multicolor bioluminescence could be the basis for using Luc as a single dual reporter gene, a bioindicator of cellular stress, and a probe for intracellular changes of pH. [13] Several hypotheses have been advanced over the years to try to explain this phenomenon. Keto-enol tautomerism, [14] rotation about a CÀC bond, [15] resonance-based structure, [16] rigidity of the active site, [17] and interaction of the emitter with a cation [18] have all been proposed, but were rejected because they cannot explain the multicolor variation.…”

Theoretical Investigation on the Origin of Yellow‐Green Firefly Bioluminescence by Time‐Dependent Density Functional Theory

“…[18,19] Access of solvent molecules to the active site is one of the main determining factors modulating bioluminescence color. [20] We used the conductor-like screening model (COSMO) [21] in the Turbomole package [22] to treat the effect of bulk water. Explicit water molecule Water324, which often was critical in predicting absorption and emission spectra, [7,14] was added to the models.…”