title: Residues at a Single Site Differentiate Animal Cryptochromes from Cyclobutane Pyrimidine Dimer Photolyases by Affecting the Proteins' Preferences for Reduced FAD.
abstract: Cryptochromes (CRYs) and photolyases belong to the cryptochrome/photolyase family (CPF). Reduced FAD is essential for photolyases to photorepair UV-induced cyclobutane pyrimidine dimers (CPDs) or 6-4 photoproducts in DNA. In Drosophila CRY (dCRY, a type I animal CRY), FAD is converted to the anionic radical but not to the reduced state upon illumination, which might induce a conformational change in the protein to relay the light signal downstream. To explore the foundation of these differences, multiple sequence alignment of 650 CPF protein sequences was performed. We identified a site facing FAD (Ala377 in Escherichia coli CPD photolyase and Val415 in dCRY), hereafter referred to as "site 377", that was distinctly conserved across these sequences: CPD photolyases often had Ala, Ser, or Asn at this site, whereas animal CRYs had Ile, Leu, or Val. The binding affinity for reduced FAD, but not the photorepair activity of E. coli photolyase, was dramatically impaired when replacing Ala377 with any of the three CRY residues. Conversely, in V415S and V415N mutants of dCRY, FAD was photoreduced to its fully reduced state after prolonged illumination, and light-dependent conformational changes of these mutants were severely inhibited. We speculate that the residues at site 377 play a key role in the different preferences of CPF proteins for reduced FAD, which differentiate animal CRYs from CPD photolyases.
authors.affiliation: Institute of Molecular Biology and Biotechnology, Anhui Normal University, 1# Beijing East Road, Wuhu, 241000, Anhui, China.|Anhui Province Key Laboratory of Active Biological Macro-Molecules, Wannan Medical College, 22# Wenchang West Road, Wuhu, 241002, Anhui, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Xu
authors.affiliation: Institute of Molecular Biology and Biotechnology, Anhui Normal University, 1# Beijing East Road, Wuhu, 241000, Anhui, China.|Anhui Province Key Laboratory of Active Biological Macro-Molecules, Wannan Medical College, 22# Wenchang West Road, Wuhu, 241002, Anhui, China.
authors.forename: Bin
authors.identifier: 0000-0002-8416-3362
authors.initials: B
authors.lastname: Wen
authors.affiliation: Institute of Molecular Biology and Biotechnology, Anhui Normal University, 1# Beijing East Road, Wuhu, 241000, Anhui, China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Institute of Molecular Biology and Biotechnology, Anhui Normal University, 1# Beijing East Road, Wuhu, 241000, Anhui, China.
authors.forename: Changqing
authors.initials: C
authors.lastname: Tian
authors.affiliation: Institute of Molecular Biology and Biotechnology, Anhui Normal University, 1# Beijing East Road, Wuhu, 241000, Anhui, China.|Anhui Province Key Laboratory of Active Biological Macro-Molecules, Wannan Medical College, 22# Wenchang West Road, Wuhu, 241002, Anhui, China.
authors.forename: Mingcai
authors.initials: M
authors.lastname: Wu
authors.affiliation: Institute of Molecular Biology and Biotechnology, Anhui Normal University, 1# Beijing East Road, Wuhu, 241000, Anhui, China.
authors.forename: Guoping
authors.initials: G
authors.lastname: Zhu
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000596:Amino Acids / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D003001:Cloning, Molecular; D017124:Conserved Sequence; D056931:Cryptochromes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004255:Deoxyribodipyrimidine Photo-Lyase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry* / Q000378:metabolism; D015870:Gene Expression; D009154:Mutation; D010084:Oxidation-Reduction; D011485:Protein Binding; D011740:Pyrimidine Dimers / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity
chemical_list: D000596:Amino Acids; D056931:Cryptochromes; D011740:Pyrimidine Dimers; D011994:Recombinant Proteins; C058406:pyrimidine-pyrimidone dimer; D005182:Flavin-Adenine Dinucleotide; D004255:Deoxyribodipyrimidine Photo-Lyase
keywords: cryptochromes; flavin; photochemistry; photolyases