Identification of a Novel Class of Photolyases as Possible Ancestors of Their Family

Abstract

UV-irradiation induces the formation of cyclobutane pyrimidine dimers (CPDs) and 6-4 photoproducts in DNA. These two types of lesions can be directly photorepaired by CPD photolyases and 6-4 photolyases, respectively. Recently, a new class of 6-4 photolyases named iron-Sulphur Bacterial Cryptochromes and Photolyases (FeS-BCPs) were found, which were considered as the ancestors of all photolyases and their homologs — cryptochromes. However, a controversy exists regarding 6-4 photoproducts only constituting ∼10-30% of the total UV-induced lesions that primordial organisms would hardly survive without a CPD repair enzyme. By extensive phylogenetic analyses, we identified a novel class of proteins, all from eubacteria. They have relatively high similarity to class I/III CPD photolyases, especially in the putative substrate-binding and FAD-binding regions. However, these proteins are shorter, and they lack the "N-terminal α/β domain" of normal photolyases. Therefore, we named them Short Photolyase-Like (SPL). Nevertheless, similar to FeS-BCPs, some of SPLs also contain four conserved cysteines, which may also coordinate an iron-sulphur cluster as FeS-BCPs. A member from Rhodococcus fascians was cloned and expressed. It was demonstrated that the protein contains a FAD cofactor and an iron-sulphur cluster, and has CPD repair activity. It was speculated that this novel class of photolyases may be the real ancestors of the cryptochrome/photolyase family.

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

“…Photolyases and their homologs cryptochromes constitute a large protein family called cryptochrome/photolyase family (CPF). Our phylogenetic work shows that CPF proteins are divided into eight main groups: class I CPD photolyases, class III CPD photolyases (including plant cryptochromes), DASHs, DASH-likes, eukaryotic 6–4 photolyases (including animal cryptochromes), class II CPD photolyases, prokaryotic 6–4 photolyases (including FeS-BCPs), and SPLs (Figure 1A ) ( 25 ). From the whole genome shotgun sequences of S. elongatus PCC 7942 ( 28 ), three CPF genes were identified: SephrA encodes a class I CPD photolyase; and SephrB and SephrC encode a FeS-BCP and a SPL, respectively (Figure 1A ).…”

“…To clarify the exact types of the proteins encoded by the hit genes, phylogenetic analysis was performed by MEGA 7.0 ( 29 ) with the three protein sequences and the other 571 sequences from 270 organisms of all life kingdoms retrieved by the same method. The sequences were divided into eight main groups: class I CPD photolyases, class III CPD photolyases, DASHs, DASH-likes, eukaryotic 6–4 photolyases, class II CPD photolyases, prokaryotic 6–4 photolyases (including FeS-BCPs), and SPLs (Figure 1A ) ( 25 ). The H6G84_07260, H6G84_09140 and H6G84_03645 encoding proteins were distributed into the class I CPD photolyase group, the prokaryotic 6–4 photolyase group, and the SPL group, respectively.…”

“…Genome sequence analysis showed that S. elongatus contains three photolyase/cryptochrome genes: a previously described CPD photolyase gene ( SephrA ), a FeS-BCP gene ( SephrB ), and a short photolyase-like (SPL) gene ( SephrC ) ( 25 ). In this study, we cloned the SephrB gene and expressed it in Escherichia coli cells.…”

A bifunctional CPD/(6-4)- photolyase from the cyanobacteria Synechococcus sp. PCC 7335 constitutes an UV-B inducible operon: new insights into photolyases phylogenetic evolution

“…Prokaryotic 6-4 photolyases were suggested as the first common ancestor of photolyases. However, as CPDs are the major UV-induced DNA damage it is not convincing that the 6-4 photo repair occurred earlier than the CPD one during evolution (Xu et al, 2021). Exhaustive phylogenetic trees do not include the recently described bifunctional CPD/(6-4)- photolyase, and its occurrence among other species was not explored.…”

“…Photoreactivation is a blue/ UV-A light-dependent mechanism used to specifically repair CPD or 6-4 PPs damages by photolyases (PHRs) (Pathak et al, 2019). These enzymes are a class of flavoproteins found in all organisms excluding placental mammals, who lost all genes encoding functional photolyases in the course of evolution (Xu et al;Banaś et al, 2020). During repair, flavin adenine dinucleotide (FAD) cofactor is fully reduced via Trp or Tyr surface transfer of electrons in a process named as photoreduction (7).…”