Searching for New Z-DNA/Z-RNA Binding Proteins Based on Structural Similarity to Experimentally Validated Zα Domain

Abstract

Z-DNA and Z-RNA are functionally important left-handed structures of nucleic acids, which play a significant role in several molecular and biological processes including DNA replication, gene expression regulation and viral nucleic acid sensing. Most proteins that have been proven to interact with Z-DNA/Z-RNA contain the so-called Zα domain, which is structurally well conserved. To date, only eight proteins with Zα domain have been described within a few organisms (including human, mouse, Danio rerio, Trypanosoma brucei and some viruses). Therefore, this paper aimed to search for new Z-DNA/Z-RNA binding proteins in the complete PDB structures database and from the AlphaFold2 protein models. A structure-based similarity search found 14 proteins with highly similar Zα domain structure in experimentally-defined proteins and 185 proteins with a putative Zα domain using the AlphaFold2 models. Structure-based alignment and molecular docking confirmed high functional conservation of amino acids involved in Z-DNA/Z-RNA, suggesting that Z-DNA/Z-RNA recognition may play an important role in a variety of cellular processes.

Keywords: Z-DNA; Z-RNA; Zα domain; bioinformatics; protein binding.

Figure 1

Schematic diagram of classical right-handed B-DNA, left-handed Z-DNA/Z-RNA, and Zα domain consisting of three α-helices and two β-strands. This domain is known to specifically interact with left-handed nucleic acids, mainly through its α-helix 3 and some amino acid residues of beta-strands.

Figure 2

Comparison of the reference Zα domain (PDB: 3f21) (upper left corner) with the experimentally solved proteins (or their corresponding domains) having significant structural similarity (structures are ordered according to their similarity score to the reference structure (HOP2 best, DsvD second best, etc.).

Figure 3

Sequence alignment is constructed from the structural superposition of the Zα domain of human ADAR1 protein (PDB: 3f21) and the 14 possible Z-DNA/Z-RNA binding proteins. The default colour of fully populated columns is light red, in addition, helices are coloured in yellow and strands in green. Letter colours correspond to the ClustalX colouring scheme.

Figure 4

Position of Zα domain in ADAR1 and within 14 newly described possible Z-DNA/Z-RNA binding proteins. The exact position of Zα and its structural homologs is always highlighted in yellow.

Figure 5

Representative molecular docking of human RPA2 Zα structural homolog to Z-DNA (A) and Z-RNA (B). Protein alpha helices are in red, beta-strands in green, coiled-coil regions in azure. Highlighting of Z-DNA/Z-RNA follows classic NDB colouring (guanines in green, cytosines in yellow).

Figure 6

STRING interaction network of newly identified human possible Z-DNA/Z-RNA binding proteins (in bold and higher letter size), together with 2 previously known human Z-DNA/Z-RNA binding proteins (ZBP1 and ADAR), and also with 50 first shell interactors. Clustering was made using MCL inflation parameter (3), the resulting five clusters are highlighted in distinct colours. Line thickness indicates the strength of data support and edges between different clusters are dotted.