The human RNA editing enzyme ADAR1 (double-stranded RNA deaminase I) deaminates adenine in pre-mRNA to yield inosine, which codes as guanine. ADAR1 has two left-handed Z-DNA binding domains, Z alpha and Z beta, at its NH(2)-terminus and preferentially binds Z-DNA, rather than B-DNA, with high binding affinity. The cocrystal structure of Z alpha(ADAR1) complexed to Z-DNA showed that one monomeric Z alpha(ADAR1) domain binds to one strand of double-stranded DNA and a second Z alpha(ADAR1) monomer binds to the opposite strand with 2-fold symmetry with respect to DNA helical axis. It remains unclear how Z alpha(ADAR1) protein specifically recognizes Z-DNA sequence in a sea of B-DNA to produce the stable Z alpha(ADAR1)-Z-DNA complex during the B-Z transition induced by Z alpha(ADAR1). In order to characterize the molecular recognition of Z-DNA by Z alpha(ADAR1), we performed circular dichroism (CD) and NMR experiments with complexes of Zalpha(ADAR1) bound to d(CGCGCG)(2) (referred to as CG6) produced at a variety of protein-to-DNA molar ratios. From this study, we identified the intermediate states of the CG6-Z alpha(ADAR1) complex and calculated their relative populations as a function of the Z alpha(ADAR1) concentration. These findings support an active B-Z transition mechanism in which the Z alpha(ADAR1) protein first binds to B-DNA and then converts it to left-handed Z-DNA, a conformation that is then stabilized by the additional binding of a second Z alpha(ADAR1) molecule.

Edited by Christian GriesingerKeywords: NMR Z-DNA Hydrogen exchange Z-DNA binding protein E3L Poxvirus a b s t r a c tThe Yaba-like disease viruses (YLDV) are members of the Yatapoxvirus family and have doublestranded DNA genomes. The E3L protein, which is essential for pathogenesis in the vaccinia virus, consists of two domains: an N-terminal Z-DNA binding domain and a C-terminal RNA binding domain. The crystal structure of the E3L orthologue of YLDV (yabZa E3L ) bound to Z-DNA revealed that the overall structure of yabZa E3L and its interaction with Z-DNA are very similar to those of hZa ADAR1 . Here we have performed NMR hydrogen exchange experiments on the complexes between yabZa E3L and d(CGCGCG) 2 with a variety of protein-to-DNA molar ratios. This study revealed that yabZa E3L could efficiently change the B-form helix of the d(CGCGCG) 2 to left-handed Z-DNA via the active-mono B-Z transition pathway like hZa ADAR1 .

Edited by Christian GriesingerKeywords: NMR Z-DNA Hydrogen exchange Z-DNA binding protein B-Z transition DNA-protein interaction a b s t r a c tThe human DNA-dependent activator of IFN-regulatory factor (DAI) protein, which activates the innate immune response in response to DNA, contains two tandem Z-DNA binding domains (Za and Zb) at the NH 2 terminus. The hZb DAI structure is similar to other Z-DNA binding proteins, although it demonstrates an unusual Z-DNA recognition. We performed NMR experiments on complexes of hZb DAI with DNA duplex, d(CGCGCG) 2 , at a variety of protein-to-DNA molar ratios. The results suggest that hZb DAI binds to Z-DNA via an active-di B-Z transition mechanism, where two hZb DAI proteins bind to B-DNA to form the hZb DAI -B-DNA complex; the B-DNA is subsequently converted to left-handed Z-DNA. This novel mechanism of DNA binding and B-Z conversion is distinct from Z-DNA binding of the human ADAR1 protein.

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