Title
Structural characterization of a novel KH-domain containing plant chloroplast endonuclease
Author
Ashok K. Rout, Himanshu Singh, Sunita Patel, Vandana Raghvan, Saurabh Gautam, R. Minda, Basuthkar J. Rao, Kandala V. R. Chary
Year
2018
Journal
Scientific Reports
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.
Instrument
J-810
Keywords
Circular dichroism, Secondary structure, Thermal stability, Thermodynamics, Biochemistry