The hSNM1 protein is a DNA 5'-exonuclease

Abstract

The human SNM1 protein is a member of a highly conserved group of proteins catalyzing the hydrolysis of nucleic acid substrates. Although overproduction is unstable in mammalian cells, we have overproduced a recombinant hSNM1 protein in an insect cell system. The protein is a single-strand 5′-exonuclease, like its yeast homolog. The enzyme utilizes either DNA or RNA substrates, requires a 5′-phosphate moiety, shows very little activity on double-strand substrates, and functions at a size consistent with a monomer. The exonuclease activity requires the conserved β-lactamase domain; site-directed mutagenesis of a conserved aspartate inactivates the exonuclease.

RAD18-dependent Recruitment of SNM1A to DNA Repair Complexes by a Ubiquitin-binding Zinc Finger

“…A murine knockout of RAD18 results in both MMC (crosslinker) and UV hypersensitivity (4,8). How crosslinking agents activate the ubiquitin ligase activity of RAD18 remains unknown.…”

“…Wild-type and RAD18-deficient HCT116 cells were grown as previously described (13). Human SNM1A cDNA has been described previously (8) and was subcloned into pEGFP-C1 vector (BglII and SalI). Mutants of the PIP box and UBZ domain were further generated based on the wild-type vector.…”

“…Third, in response to DNA damage, SNM1A assembles in subnuclear foci (7), further supporting its role in the DNA damage response. Fourth, SNM1A can function as a DNA exonuclease in vitro, suggesting that it may have a direct role in excising DNA ICLs (3,8).DNA repair proteins are often recruited to specific sites of DNA damage through protein-protein interaction domains. For example, the FHA domain of NBS1 and the BRCT domains of BRCA1 are required for recruitment of these DNA repair proteins to complexes containing phosphopeptide motifs (9).…”

“…Third, in response to DNA damage, SNM1A assembles in subnuclear foci (7), further supporting its role in the DNA damage response. Fourth, SNM1A can function as a DNA exonuclease in vitro, suggesting that it may have a direct role in excising DNA ICLs (3,8).…”

Fan1 deficiency results in DNA interstrand cross-link repair defects, enhanced tissue karyomegaly, and organ dysfunction

“…The in vitro activity of FAN1 on substrates containing an ICL overlaps with the activity of SNM1A, one of the three human homologs of Pso2, a nuclease that functions in ICL repair in Saccharomyces cerevisiae (Henriques and Moustacchi 1980;Ruhland et al 1981;Hejna et al 2007;Wang et al 2011). Mammalian SNM1A shares the most similarity with Pso2 and is the only homolog that can complement the ICL sensitivity of pso2Δ yeast (Hazrati et al 2008).…”