title: Single-Stranded Nucleic Acids Bind to the Tetramer Interface of SAMHD1 and Prevent Formation of the Catalytic Homotetramer.
abstract: Sterile alpha motif and HD domain protein 1 (SAMHD1) is a unique enzyme that plays important roles in nucleic acid metabolism, viral restriction, and the pathogenesis of autoimmune diseases and cancer. Although much attention has been focused on its dNTP triphosphohydrolase activity in viral restriction and disease, SAMHD1 also binds to single-stranded RNA and DNA. Here we utilize a UV cross-linking method using 5-bromodeoxyuridine-substituted oligonucleotides coupled with high-resolution mass spectrometry to identify the binding site for single-stranded nucleic acids (ssNAs) on SAMHD1. Mapping cross-linked amino acids on the surface of existing crystal structures demonstrated that the ssNA binding site lies largely along the dimer-dimer interface, sterically blocking the formation of the homotetramer required for dNTPase activity. Surprisingly, the disordered C-terminus of SAMHD1 (residues 583-626) was also implicated in ssNA binding. An interaction between this region and ssNA was confirmed in binding studies using the purified SAMHD1 583-626 peptide. Despite a recent report that SAMHD1 possesses polyribonucleotide phosphorylase activity, we did not detect any such activity in the presence of inorganic phosphate, indicating that nucleic acid binding is unrelated to this proposed activity. These data suggest an antagonistic regulatory mechanism in which the mutually exclusive oligomeric state requirements for ssNA binding and dNTP hydrolase activity modulate these two functions of SAMHD1 within the cell.
authors: Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine , 725 North Wolfe Street, WBSB 314, Baltimore, Maryland 21205, United States.
authors: Kyle J
authors: KJ
authors: Seamon
authors: Department of Medicine, Division of Clinical Pharmacology, Johns Hopkins University School of Medicine , 725 North Wolfe Street, Biophysics 307, Baltimore, Maryland 21205, United States.
authors: Namandjé N
authors: NN
authors: Bumpus
authors: Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine , 725 North Wolfe Street, WBSB 314, Baltimore, Maryland 21205, United States.
authors: James T
authors: JT
authors: Stivers
mesh_terms: D001704:Biopolymers / Q000737:chemistry*; D001973:Bromodeoxyuridine / Q000737:chemistry; D002384:Catalysis; D002853:Chromatography, Liquid; D003001:Cloning, Molecular; D004277:DNA, Single-Stranded / Q000737:chemistry*; D006801:Humans; D013058:Mass Spectrometry; D020559:Monomeric GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D012313:RNA / Q000737:chemistry*; D000076106:SAM Domain and HD Domain-Containing Protein 1
chemical_list: D001704:Biopolymers; D004277:DNA, Single-Stranded; D012313:RNA; D000076106:SAM Domain and HD Domain-Containing Protein 1; C484280:SAMHD1 protein, human; D020559:Monomeric GTP-Binding Proteins; D001973:Bromodeoxyuridine