Light activation of the LOV protein vivid generates a rapidly exchanging dimer.
The fungal photoreceptor Vivid (VVD) plays an important role in the adaptation of blue-light responses in Neurospora crassa. VVD, an FAD-binding LOV (light, oxygen, voltage) protein, couples light-induced cysteinyl adduct formation at the flavin ring to conformational changes in the N-terminal cap (Ncap) of the VVD PAS domain. Size-exclusion chromatography (SEC), equilibrium ultracentrifugation, and static and dynamic light scattering show that these conformational changes generate a rapidly exchanging VVD dimer, with an expanded hydrodynamic radius. A three-residue N-terminal beta-turn that assumes two different conformations in a crystal structure of a VVD C71V variant is essential for light-state dimerization. Residue substitutions at a critical hinge between the Ncap and PAS core can inhibit or enhance dimerization, whereas a Tyr to Trp substitution at the Ncap-PAS interface stabilizes the light-state dimer. Cross-linking through engineered disulfides indicates that the light-state dimer differs considerably from the dark-state dimer found in VVD crystal structures. These results verify the role of Ncap conformational changes in gating the photic response of N. crassa and indicate that LOV-LOV homo- or heterodimerization may be a mechanism for regulating light-activated gene expression.
Zoltowski
Brian D
BD
Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853, USA.
Crane
Brian R
BR
D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D018360:Crystallography, X-Ray; D019281:Dimerization; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism* / Q000528:radiation effects*; D007700:Kinetics; D008027:Light; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D009492:Neurospora crassa / Q000378:metabolism* / Q000528:radiation effects*; D017433:Protein Structure, Secondary; D016415:Sequence Alignment
D005656:Fungal Proteins; D050505:Mutant Proteins; C423283:VVD protein, Neurospora crassa