Structural basis of cytokinin transport by the Arabidopsis AZG2

Cytokinins are adenine derivatives serving as phytohormones, which are essential regulators of plant growth, development, and response to environmental factors. The transport process of cytokinins determines their spatial distributions and is critical to their functions. The AZA-GUANINE RESISTANT (AZG) family member AZG2 in Arabidopsis thaliana has been identified as a cytokinin and purine transporter. Here, we characterize the binding and transport of AZG2 towards the natural cytokinin, trans-zeatin (tZ), as well as adenine. AZG2 structures are determined in both the substrate-unbound, adenine-bound, and tZ-bound states, under both acidic and neutral pH. Key residues involved in substrate binding are identified. Two distinct conformations are observed in the tZ-bound state of AZG2 in the neutral pH. Structural analysis reveals the structural dynamics of AZG2 during cytokinin transport, which fit into the elevator-type transport model. These results provide insights into the molecular mechanism of cytokinin transport in plants. This study characterized the substrate binding and transport mechanisms of the plant purine/cytokinin transporter AZG2. Conformational dynamics were examined by determining AZG2 structures in the apo or substrate-bound states under different pH.