Structural refinement of a key tryptophan residue in the BLUF photoreceptor AppA by ultraviolet resonance Raman spectroscopy.
The flavin-adenine-dinucleotide-binding BLUF domain constitutes a new class of blue-light receptors, and the N-terminal domain of AppA is a representative of this family. The BLUF domain is of special interest because it uses a rigid flavin rather than an isomerizable chromophore, such as a rhodopsin or phytochrome, for its light-activation process. Crystal and solution structures of several BLUF domains were recently obtained, and their overall structures are consistent. However, there is a key ambiguity regarding the position of a conserved tryptophan (Trp-104 in AppA), in that this residue was found either close to flavin (Trp(in) conformation) or exposed to the solvent (Trp(out) conformation). The location of Trp-104 is a crucial factor in understanding the photocycle mechanism of BLUF domains, because this residue has been shown to play an essential role in the activation of AppA. In this study, we demonstrated a Trp(in) conformation for the BLUF domain of AppA through direct observation of the vibrational spectrum of Trp-104 by ultraviolet resonance Raman spectroscopy, and also observed light-induced conformational and environmental changes in Trp-104. This study provides a structural basis for future investigations of the photocycle mechanism of BLUF proteins.
Unno
Masashi
M
Department of Chemistry and Applied Chemistry, Faculty of Science and Engineering, Saga University, Saga, Japan. unno@cc.saga-u.ac.jp
Kikuchi
Sadato
S
Masuda
Shinji
S
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003624:Darkness; D005420:Flavoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D009154:Mutation; D017434:Protein Structure, Tertiary / Q000528:radiation effects; D012996:Solutions; D013059:Spectrum Analysis, Raman; D014364:Tryptophan; D014466:Ultraviolet Rays
C116785:AppA protein, Rhodobacter sphaeroides; D001426:Bacterial Proteins; D005420:Flavoproteins; D012996:Solutions; D014364:Tryptophan