STRUCTURE OF BACTERIAL LUCIFERASE BETA(2) HOMODIMER - IMPLICATIONS FOR FLAVIN BINDING

The crystal structure of the beta(2) homodimer of Vibrio harveyi lucif erase has been determined to 2.5 Angstrom resolution by molecular repl acement. Crystals were grown serendipitously using the alpha beta form of the enzyme. The subunits of the homodimer share considerable struc tural homology to the beta subunit of the alpha beta luciferase hetero dimer. The four C-terminal residues that are disordered in the alpha b eta structure are fully resolved in our structure. Four peptide bonds have been flipped relative to their orientations in the beta subunit o f the alpha beta structure. The dimer interface of the homodimer is sm aller than the interface of the heterodimer in terms of buried surface area and number of hydrogen bonds and salt links. Inspection of the s ubunits of our structure suggests that FMNH(2) cannot bind to the beta (2) enzyme at the site that has been proposed for the alpha beta enzym e. However, we do uncover a potential FMNH(2) binding pocket in the di mer interface, and we model FMN into this site. This proposed flavin b inding motif is consistent with several lines of biochemical and struc tural evidence and leads to several conclusions. First, only one FMNH( 2) binds per homodimer. Second, we predict that reduced FAD and ribofl avin should be poor substrates for beta(2). Third, the reduced activit y of beta(2) compared to alpha beta is due to solvent exposure of the isoalloxazine ring in the beta(2) active site. Finally, we raise the q uestion of whether our proposed flavin binding site could also be the binding site for flavin in the alpha beta enzyme.