The Arg-62 Residues of the TREX1 Exonuclease Act Across the Dimer Interface Contributing to Catalysis in the Opposing Protomers [Enzymology]

March 9th, 2014 by Fye, J. M., Coffin, S. R., Orebaugh, C. D., Hollis, T., Perrino, F. W.

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 forty 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-Goutieres 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 TREX1R62A/R62A homodimer exhibits approximately 50-fold reduced ssDNA and dsDNA degradation activities relative to TREX1WT. 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 TREX1D18H/R62A, TREX1D18N/R62A, TREX1D200H/R62A, and TREX1D200N/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.