Thermodynamic analysis of Za domain-nucleic acid interactions

DNA/RNA molecules adopting the left-handed conformation (Z-form) have been attributed with immunogenic properties. However, their biological role and importance have been a topic of debate for many years. The discovery of Z-DNA/RNA binding domains (Z alpha domains) in varied proteins that are involved in the innate immune response, such as the interferon inducible form of the RNA editing enzyme ADAR1 (p150), Z-DNA binding protein 1 (ZBP1), the fish kinase PKZ and the poxvirus inhibitor of interferon response E3L, indicates important roles of Z-DNA/RNA in immunity and self/non-self-discrimin-ation. Such Z alpha domain-containing proteins recognize left-handed Z-DNA/RNA in a con-formation-specific manner. Recent studies have implicated these domains in virus recognition. Given these important emerging roles for the Z alpha domains, it is pivotal to understand the mechanism of recognition of the Z-DNA/Z-RNA by these domains. To this end, we assessed the binding thermodynamics of Z alpha domain from ORF112 and ADAR1 on T(CG)3 and T(CG)6 oligonucleotides which have high propensity to adopt the Z-conformation. Our study highlights important differences in the mode of oligonucleo-tide binding by the two Z alpha domains originating from different proteins. Site-directed mutagenesis was employed together with isothermal titration calorimetry to tease apart finer details of the binding thermodynamics. Our work advances the understanding on binding thermodynamics of Z alpha domains to their cognate nucleic acid substrates and paves the ground for future efforts to gain a complete appreciation of this process.

Automated summary

DNA/RNA molecules adopting the left-handed conformation (Z-form) have immunogenic properties and play important roles in immunity and self/non-self-discrimination. Z alpha domains in different proteins recognize and bind left-handed Z-DNA/RNA in a conformation-specific manner, and have been implicated in virus recognition.