Abstract
Human EXOG (hEXOG) is a 5′-exonuclease that is crucial for mitochondrial DNA repair; the enzyme belongs to a nonspecific nuclease family that includes the apoptotic endonuclease EndoG. Here we report biochemical and structural studies of hEXOG, including structures in its apo form and in a complex with DNA at 1.81 and 1.85 Å resolution, respectively. A Wing domain, absent in other ββα-Me members, suppresses endonuclease activity, but confers on hEXOG a strong 5′-dsDNA exonuclease activity that precisely excises a dinucleotide using an intrinsic ‘tape-measure’. The symmetrical apo hEXOG homodimer becomes asymmetrical upon binding to DNA, providing a structural basis for how substrate DNA bound to one active site allosterically regulates the activity of the other. These properties of hEXOG suggest a pathway for mitochondrial BER that provides an optimal substrate for subsequent gap-filling synthesis by DNA polymerase γ.
Highlights
Human EXOG is a 50-exonuclease that is crucial for mitochondrial DNA repair; the enzyme belongs to a nonspecific nuclease family that includes the apoptotic endonuclease EndoG
HEXOG is found in a complex with other mitochondrial DNA (mtDNA) repair enzymes, APE1, Pol g and ligase III; their interaction is enhanced by oxidative stress, and is regulated by poly(ADP-ribose) polymerase-117
Our studies suggest that mtDNA repair is performed by long-patch Base excision repair (BER) that provides an optimal substrate for Pol g when fulfilling its role in DNA repair
Summary
Human EXOG (hEXOG) is a 50-exonuclease that is crucial for mitochondrial DNA repair; the enzyme belongs to a nonspecific nuclease family that includes the apoptotic endonuclease EndoG. The symmetrical apo hEXOG homodimer becomes asymmetrical upon binding to DNA, providing a structural basis for how substrate DNA bound to one active site allosterically regulates the activity of the other These properties of hEXOG suggest a pathway for mitochondrial BER that provides an optimal substrate for subsequent gap-filling synthesis by DNA polymerase g. Base excision repair (BER) is the major mechanism for correcting mtDNA oxidative damage[6] In this multi-enzyme reaction pathway, the DNA product from the previous reaction is the substrate for the [7]. Ectopic expression of hEXOG increases resistance to oxidative stress of proliferating myoblasts[15], and depletion of EXOG increases oxidative consumption rate in primary neonatal rat ventricular cardiomyocytes[16] These studies provide strong support to the idea that hEXOG is involved in mtDNA repair. HEXOG is found in a complex with other mtDNA repair enzymes, APE1, Pol g and ligase III; their interaction is enhanced by oxidative stress, and is regulated by poly(ADP-ribose) polymerase-117
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