Backbone Dynamics Study of the Zβ Domain of Human DAI Bound to Z‐DNA#

Abstract

Innate immune response is an essential defense against infections. Human DNA-dependent activator of IFN-regulatory factor (DAI; also known as ZBP1 or DLM-1) protein activates the innate immune response in response to DNA. The DAI protein contains two tandem Z-DNA binding domains (Zα and Zβ) at the N-terminus. The two Z-DNA binding domains are thought to be essential for full activation of a DNA-dependent immune response in vivo. The crystal structure of the Zα domain of human ADAR1 (ZαADAR1) in complex with Z-DNA shows that two Zα domains bind to each strand of double-stranded DNA and have twofold symmetry with respect to the DNA's helical axis. Structural studies with the Zβ domain of human DAI (ZβDAI) in complex with Z-DNA suggest that the ZβDAI structure is similar to that of ZαADAR1, although it demonstrates an unusual Z-DNA recognition. A previous NMR study of the ZβDAI complexed with a six-base-pair (6-bp) DNA duplex, d(CG)3, suggests that ZβDAI binds to Z-DNA via an activedi B–Z transition mechanism, where two ZβDAI proteins bind to B-DNA to form the ZβDAI–B-DNA complex; the B-DNA is subsequently converted to left-handed Z-DNA. To investigate the structural and dynamic properties of ZβDAI when it binds toDNAduplex and induces the B–Z transition in a DNA duplex, we have performed NMR backbone dynamics experiments on the free ZβDAI and the ZβDAI–d (CG)3 complex. The results revealed that ZβDAI exhibits a distinct change in the dynamics during theB–Z transition ofDNA duplex compared to ZαADAR1. This study provides valuable insights into the molecular mechanism of the B–Z transition induced by ZβDAI.