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Characterization of the Human SNM1A and SNM1B/Apollo DNA Repair Exonucleases
- Source :
- The Journal of Biological Chemistry
- Publication Year :
- 2012
- Publisher :
- Elsevier BV, 2012.
-
Abstract
- Background: The nucleases hSNM1A and hSNM1B are implicated in DNA interstrand cross-link repair. Results: hSNM1A and hSNM1B were biochemically characterized using undamaged and cross-linked DNA. A real-time assay for the nucleases suitable for inhibitor identification was developed. Conclusion: Preferential hSNM1A activation by high molecular weight and cross-linked DNA was observed. Significance: This work provides a basis for hSNM1A inhibitor development for improved cancer therapy.<br />Human SNM1A and SNM1B/Apollo have both been implicated in the repair of DNA interstrand cross-links (ICLs) by cellular studies, and SNM1B is also required for telomere protection. Here, we describe studies on the biochemical characterization of the SNM1A and SNM1B proteins. The results reveal some fundamental differences in the mechanisms of the two proteins. Both SNM1A and SNM1B digest double-stranded and single-stranded DNA with a 5′-to-3′ directionality in a reaction that is stimulated by divalent cations, and both nucleases are inhibited by the zinc chelator o-phenanthroline. We find that SNM1A has greater affinity for single-stranded DNA over double-stranded DNA that is not observed with SNM1B. Although both proteins demonstrate a low level of processivity on low molecular weight DNA oligonucleotide substrates, when presented with high molecular weight DNA, SNM1A alone is rendered much more active, being capable of digesting kilobase-long stretches of DNA. Both proteins can digest past ICLs induced by the non-distorting minor groove cross-linking agent SJG-136, albeit with SNM1A showing a greater capacity to achieve this. This is consistent with the proposal that SNM1A and SNM1B might exhibit some redundancy in ICL repair. Together, our work establishes differences in the substrate selectivities of SNM1A and SNM1B that are likely to be relevant to their in vivo roles and which might be exploited in the development of selective inhibitors.
- Subjects :
- DNA Repair
DNA repair
DNA damage
Deoxyribozyme
DNA, Single-Stranded
Cell Cycle Proteins
Biology
Biochemistry
DNA-binding protein
Substrate Specificity
DNA Enzymes
DNase
chemistry.chemical_compound
Metalloproteins
Escherichia coli
Humans
Magnesium
Protein–DNA interaction
DNA Cleavage
Enzyme Inhibitors
Molecular Biology
Chelating Agents
Enzyme Assays
Fluorescent Dyes
Oligonucleotide
Hydrolysis
Nuclear Proteins
Metallo-β-lactamase
DNA
Cell Biology
Processivity
Hydrogen-Ion Concentration
Recombinant Proteins
DNA-Binding Proteins
DNA Interstrand Cross-link
DNA Repair Enzymes
Exodeoxyribonucleases
chemistry
Enzymology
Nucleic Acid Enzymology
RNA
Fluorescein
DNA Damage
Plasmids
Protein Binding
Subjects
Details
- ISSN :
- 00219258
- Volume :
- 287
- Database :
- OpenAIRE
- Journal :
- Journal of Biological Chemistry
- Accession number :
- edsair.doi.dedup.....bc8bd54f12273d4f45530a2f69f547ad