Reverse structural genomics: an unusual flavin-binding site in a putative protease from Bacteroides thetaiotaomicron.
The structure of a putative protease from Bacteroides thetaiotaomicron features an unprecedented binding site for flavin mononucleotide. The flavin isoalloxazine ring is sandwiched between two tryptophan residues in the interface of the dimeric protein. We characterized the recombinant protein with regard to its affinity for naturally occurring flavin derivatives and several chemically modified flavin analogs. Dissociation constants were determined by isothermal titration calorimetry. The protein has high affinity to naturally occurring flavin derivatives, such as riboflavin, FMN, and FAD, as well as lumichrome, a photodegradation product of flavins. Similarly, chemically modified flavin analogs showed high affinity to the protein in the nanomolar range. Replacement of the tryptophan by phenylalanine gave rise to much weaker binding, whereas in the tryptophan to alanine variant, flavin binding was abolished. We propose that the protein is an unspecific scavenger of flavin compounds and may serve as a storage protein in vivo.
Institute of Biochemistry, Graz University of Technology, A-8010 Graz, Austria.
Tanja
T
Knaus
Elisabeth
E
Eger
Julia
J
Koop
Steve
S
Stipsits
Cynthia L
CL
Kinsland
Steven E
SE
Ealick
Peter
P
Macheroux
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001439:Bacteroides / Q000201:enzymology* / Q000235:genetics; D005486:Flavin Mononucleotide / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D023281:Genomics; D010447:Peptide Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012256:Riboflavin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D005182:Flavin-Adenine Dinucleotide; D005486:Flavin Mononucleotide; D010447:Peptide Hydrolases; D012256:Riboflavin