Comparative photochemistry of animal type 1 and type 4 cryptochromes.
Cryptochromes (CRYs) are blue-light photoreceptors with known or presumed functions in light-dependent and light-independent gene regulation in plants and animals. Although the photochemistry of plant CRYs has been studied in some detail, the photochemical behavior of animal cryptochromes remains poorly defined in part because it has been difficult to purify animal CRYs with their flavin cofactors. Here we describe the purification of type 4 CRYs of zebrafish and chicken as recombinant proteins with full flavin complement and compare the spectroscopic properties of type 4 and type 1 CRYs. In addition, we analyzed photoinduced proteolytic degradation of both types of CRYs in vivo in heterologous systems. We find that even though both types of CRYs contain stoichiometric flavin, type 1 CRY is proteolytically degraded by a light-initiated reaction in Drosophila S2, zebrafish Z3, and human HEK293T cell lines, but zebrafish CRY4 (type 4) is not. In vivo degradation of type 1 CRYs does not require continuous illumination, and a single light flash of 1 ms duration leads to degradation of about 80% of Drosophila CRY in 60 min. Finally, we demonstrate that in contrast to animal type 2 CRYs and Arabidopsis CRY1 neither insect type 1 nor type 4 CRYs have autokinase activities.
Ozturk
Nuri
N
Department of Biochemistry and Biophysics, University of North Carolina School of Medicine, Chapel Hill, North Carolina 27599, USA.
Selby
Christopher P
CP
Song
Sang-Hun
SH
Ye
Rui
R
Tan
Chuang
C
Kao
Ya-Ting
YT
Zhong
Dongping
D
Sancar
Aziz
A
D000818:Animals; D000852:Anopheles; D030161:Avian Proteins / Q000737:chemistry* / Q000378:metabolism; D002460:Cell Line; D002645:Chickens; D056931:Cryptochromes; D004255:Deoxyribodipyrimidine Photo-Lyase / Q000737:chemistry; D029721:Drosophila Proteins / Q000737:chemistry / Q000378:metabolism / Q000528:radiation effects; D004926:Escherichia coli / Q000201:enzymology; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry; D005415:Flavins / Q000737:chemistry* / Q000378:metabolism / Q000528:radiation effects; D005420:Flavoproteins / Q000737:chemistry* / Q000378:metabolism / Q000528:radiation effects; D006801:Humans; D008027:Light; D010777:Photochemistry / Q000379:methods; D051381:Rats; D029961:Zebrafish Proteins / Q000737:chemistry* / Q000378:metabolism
D030161:Avian Proteins; C538963:CRY1 protein, human; C538965:Cry1 protein, rat; D056931:Cryptochromes; D029721:Drosophila Proteins; D005415:Flavins; D005420:Flavoproteins; D029961:Zebrafish Proteins; D005182:Flavin-Adenine Dinucleotide; D004255:Deoxyribodipyrimidine Photo-Lyase