Structural characterization of a novel KH-domain containing plant chloroplast endonuclease

Abstract

Chlamydomonas reinhardtii is a single celled alga that undergoes apoptosis in response to UV-C irradiation. UVI31+, a novel UV-inducible DNA endonuclease in C. reinhardtii, which normally localizes near cell wall and pyrenoid regions, gets redistributed into punctate foci within the whole chloroplast, away from the pyrenoid, upon UV-stress. Solution NMR structure of the first putative UV inducible endonuclease UVI31+ revealed an α1–β1–β2–α2–α3–β3 fold similar to BolA and type II KH-domain ubiquitous protein families. Three α−helices of UVI31+ constitute one side of the protein surface, which are packed to the other side, made of three-stranded β–sheet, with intervening hydrophobic residues. A twenty-three residues long polypeptide stretch (D54-H76) connecting β1 and β2 strands is found to be highly flexible. Interestingly, UVI31+ recognizes the DNA primarily through its β–sheet. We propose that the catalytic triad residues involving Ser114, His95 and Thr116 facilitate DNA endonuclease activity of UVI31+. Further, decreased endonuclease activity of the S114A mutant is consistent with the direct participation of Ser114 in the catalysis. This study provides the first structural description of a plant chloroplast endonuclease that is regulated by UV-stress response.