title: The Arg-62 residues of the TREX1 exonuclease act across the dimer interface contributing to catalysis in the opposing protomers.
abstract: TREX1 is a 3'-deoxyribonuclease that degrades single- and double-stranded DNA (ssDNA and dsDNA) to prevent inappropriate nucleic acid-mediated immune activation. More than 40 different disease-causing TREX1 mutations have been identified exhibiting dominant and recessive genetic phenotypes in a spectrum of autoimmune disorders. Mutations in TREX1 at positions Asp-18 and Asp-200 to His and Asn exhibit dominant autoimmune phenotypes associated with the clinical disorders familial chilblain lupus and Aicardi-Goutières syndrome. Our previous biochemical studies showed that the TREX1 dominant autoimmune disease phenotype depends upon an intact DNA-binding process coupled with dysfunctional active site chemistry. Studies here show that the TREX1 Arg-62 residues extend across the dimer interface into the active site of the opposing protomer to coordinate substrate DNA and to affect catalysis in the opposing protomer. The TREX1(R62A/R62A) homodimer exhibits ∼50-fold reduced ssDNA and dsDNA degradation activities relative to TREX1(WT). The TREX1 D18H, D18N, D200H, and D200N dominant mutant enzymes were prepared as compound heterodimers with the TREX1 R62A substitution in the opposing protomer. The TREX1(D18H/R62A), TREX1(D18N/R62A), TREX1(D200H/R62A), and TREX1(D200N/R62A) compound heterodimers exhibit higher levels of ss- and dsDNA degradation activities than the homodimers demonstrating the requirement for TREX1 Arg-62 residues to provide necessary structural elements for full catalytic activity in the opposing TREX1 protomer. This concept is further supported by the loss of dominant negative effects in the TREX1 D18H, D18N, D200H, and D200N compound heterodimers. These data provide compelling evidence for the required TREX1 dimeric structure for full catalytic function.
authors.affiliation: Department of Biochemistry, Wake Forest School of Medicine, Winston-Salem, North Carolina 27157.
authors.forename: Jason M
authors.initials: JM
authors.lastname: Fye
authors.affiliation: Department of Biochemistry, Wake Forest School of Medicine, Winston-Salem, North Carolina 27157.
authors.forename: Stephanie R
authors.initials: SR
authors.lastname: Coffin
authors.affiliation: Department of Biochemistry, Wake Forest School of Medicine, Winston-Salem, North Carolina 27157.
authors.forename: Clinton D
authors.initials: CD
authors.lastname: Orebaugh
authors.affiliation: Department of Biochemistry, Wake Forest School of Medicine, Winston-Salem, North Carolina 27157.
authors.forename: Thomas
authors.initials: T
authors.lastname: Hollis
authors.affiliation: Department of Biochemistry, Wake Forest School of Medicine, Winston-Salem, North Carolina 27157. Electronic address: fperrino@wakehealth.edu.
authors.forename: Fred W
authors.initials: FW
authors.lastname: Perrino
mesh_terms: D019943:Amino Acid Substitution; D001120:Arginine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020274:Autoimmune Diseases of the Nervous System / Q000201:enzymology / Q000235:genetics; D002384:Catalysis; D004277:DNA, Single-Stranded / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005090:Exodeoxyribonucleases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D020125:Mutation, Missense; D009421:Nervous System Malformations / Q000201:enzymology / Q000235:genetics; D010750:Phosphoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D055503:Protein Multimerization / Q000502:physiology*; D020836:Protein Structure, Quaternary; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D004277:DNA, Single-Stranded; D010750:Phosphoproteins; D021122:Protein Subunits; D001120:Arginine; D005090:Exodeoxyribonucleases; C429526:three prime repair exonuclease 1
keywords: Autoimmune Diseases; DNA; DNase; Dimer; Enzyme Mechanisms; Exonuclease; Nucleic Acid; TREX1