Identification and characterization of a prokaryotic 6-4 photolyase from Synechococcus elongatus with a deazariboflavin antenna chromophore

Abstract

Synechococcus elongatus, formerly known as Anacystis nidulans, is a representative species of cyanobacteria. It is also a model organism for the study of photoreactivation, which can be fully photoreactivated even after receiving high UV doses. However, for a long time, only one photolyase was found in S. elongatus that is only able to photorepair UV induced cyclobutane pyrimidine dimers (CPDs) in DNA. Here, we characterize another photolyase in S. elongatus, which belongs to iron-sulfur bacterial cryptochromes and photolyases (FeS-BCP), a subtype of prokaryotic 6–4 photolyases. This photolyase was named SePhrB that could efficiently photorepair 6–4 photoproducts in DNA. Chemical analyses revealed that SePhrB contains a catalytic FAD cofactor and an iron-sulfur cluster. All of previously reported FeS-BCPs contain 6,7-dimethyl-8-ribityllumazine (DMRL) as their antenna chromophores. Here, we first demonstrated that SePhrB possesses 7,8-didemethyl-8-hydroxy-5-deazariboflavin (8-HDF) as an antenna chromophore. Nevertheless, SePhrB could be photoreduced without external electron donors. After being photoreduced, the reduced FAD cofactor in SePhrB was extremely stable against air oxidation. These results suggest that FeS-BCPs are more diverse than expected which deserve further investigation.

A deazariboflavin chromophore kinetically stabilizes reduced FAD state in a bifunctional cryptochrome

“…( c ) Midpoint potentials of the FAD ox /FADH − redox pair in (6–4)PP-repairing proteins were ~ − 290 mV vs. SHE. The value was considerably lower than those reported for CPD PL derived from Synechococcus elongatus ( Se PhrA), formerly called Anacystis nidulans CPD PL 11 , 24 and for the solution FAD (aq) state 33 . Error bars for our samples reflect the standard deviation for n = 3.…”

Crystal structure of Synechococcus elongatus 6-4 photolyase suggests a novel repairing mechanism

“…They are involved in structure folding, electron transferring, and/or substrate binding of many proteins 13 . Recently, an increasing number of nucleic acid processing enzymes that contain iron-sulfur clusters were discovered 23 , including eukaryotic primase 24 , prokaryotic 6-4 photolyases [6][7][8][9][10] and SPLs 14 . As many prokaryotic 6-4 photolyases 7-10 , SePhrB also contains a [4Fe-4S] cluster, which is coordinated by four conserved cysteine residues: Cys346, Cys434, Cys437, and Cys450, in a roof-like region at Cterminus (Fig.…”

“…A CPD photolyase from Synechococcus elongatus (also known as Anacystis nidulans) has been extensively studied [3][4][5] . Recently, we identified a prokaryotic 6-4 photolyase from S. elongatus (SePhrB) 6 . Crystal structure of SePhrB present here shows that the protein has three cofactors: an iron-sulfur cluster; a 7,8-didemethyl-8-hydroxy-5-deazariboflavin (8-HDF); and a FAD.…”

Insights into Molecular Structure of Pterins Suitable for Biomedical Applications

“…In CPF proteins, in addition to the main FAD chromophore, there are pigments (derivatives of flavins and pterins) that act as a “light-harvesting antenna” ( Figure 23 ). Five molecules have been described as antennas of CPF [ 233 ]: 5,10-methenyl-5,6,7,8-tetrahydrofolate (MTHF) acts as an antenna in most eukaryotes and some prokaryotes [ 245 , 246 ]; 7-desmethyl-8-hydroxy-5-deazariboflavin (8-HDF)—in some prokaryotes and protozoa eukaryotes, which have a biosynthetic pathway for this compound [ 247 ]; FMN or the second FAD can also function as an antenna [ 248 ]; it has recently been shown that 6,7-dimethyl-8-ribityllumazine can function as an antenna in some prokaryotic (6-4)-photolyases ( Figure 23 ) [ 249 ]. …”

“…7-desmethyl-8-hydroxy-5-deazariboflavin (8-HDF)—in some prokaryotes and protozoa eukaryotes, which have a biosynthetic pathway for this compound [ 247 ];…”