Effects of Conserved Wedge Domain Residues on DNA Binding Activity of Deinococcus radiodurans RecG Helicase

Sun-Wook Jeong; Min-Kyu Kim; Lei Zhao; Seul-Ki Yang; Jong-Hyun Jung; Heon-Man Lim; Sangyong Lim

doi:10.3389/fgene.2021.634615

Effects of Conserved Wedge Domain Residues on DNA Binding Activity of Deinococcus radiodurans RecG Helicase

Front Genet. 2021 Feb 4:12:634615. doi: 10.3389/fgene.2021.634615. eCollection 2021.

Authors

Sun-Wook Jeong^{1

2}, Min-Kyu Kim¹, Lei Zhao¹, Seul-Ki Yang¹, Jong-Hyun Jung^{1

3}, Heon-Man Lim², Sangyong Lim^{1

3}

Affiliations

¹ Radiation Research Division, Korea Atomic Energy Research Institute, Jeongeup, South Korea.
² Department of Biological Sciences, College of Biological Sciences and Biotechnology, Chungnam National University, Daejeon, South Korea.
³ Department of Radiation Science and Technology, University of Science and Technology, Daejeon, South Korea.

Abstract

Deinococcus radiodurans is extremely resistant to ionizing radiation and has an exceptional ability to repair DNA damage caused by various DNA-damaging agents. D. radiodurans uses the same DNA-repair strategies as other prokaryotes, but certain proteins involved in the classical DNA repair machinery have characteristics different from their counterparts. RecG helicase, which unwinds a variety of branched DNA molecules, such as Holliday junctions (HJ) and D-loops, plays important roles in DNA repair, recombination, and replication. Primary sequence analysis of RecG from a number of bacterial species revealed that three amino acids (QPW) in the DNA-binding wedge domain (WD) are well-conserved across the Deinococcus RecG proteins. Interactions involving these conserved residues and DNA substrates were predicted in modeled domain structures of D. radiodurans RecG (DrRecG). Compared to the WD of Escherichia coli RecG protein (EcRecG) containing FSA amino acids corresponding to QPW in DrRecG, the HJ binding activity of DrRecG-WD was higher than that of EcRecG-WD. Reciprocal substitution of FSA and QPW increased and decreased the HJ binding activity of the mutant WDs, EcRecG-WD_QPW, and DrRecG-WD_FSA, respectively. Following γ-irradiation treatment, the reduced survival rate of DrRecG mutants (ΔrecG) was fully restored by the expression of DrRecG, but not by that of EcRecG. EcRecG_QPW also enhanced γ-radioresistance of ΔrecG, whereas DrRecG_FSA did not. ΔrecG cells complemented in trans by DrRecG and EcRecG_QPW reconstituted an intact genome within 3 h post-irradiation, as did the wild-type strain, but ΔrecG with EcRecG and DrRecG_FSA exhibited a delay in assembly of chromosomal fragments induced by γ-irradiation. These results suggested that the QPW residues facilitate the association of DrRecG with DNA junctions, thereby enhancing the DNA repair efficiency of DrRecG.

Keywords: DNA repair; Deinococcus radiodurans; RecG helicase; radiation resistance; wedge domain.