calmodulin-like protein CML36 is a calcium (Ca) sensor that interacts with the plasma membrane Ca-ATPase isoform ACA8 and stimulates its activity.

Calmodulin-like (CML) proteins are major EF-hand-containing, calcium (Ca)-binding proteins with crucial roles in plant development and in coordinating plant stress tolerance. Given their abundance in plants, the properties of Ca sensors and identification of novel target proteins of CMLs deserve special attention. To this end, we recombinantly produced and biochemically characterized CML36 from We analyzed Ca and Mg binding to the individual EF-hands, observed metal-induced conformational changes, and identified a physiologically relevant target. CML36 possesses two high-affinity Ca/Mg mixed binding sites and two low-affinity Ca-specific sites. Binding of Ca induced an increase in the α-helical content and a conformational change that lead to the exposure of hydrophobic regions responsible for target protein recognition. Cation binding, either Ca or Mg, stabilized the secondary and tertiary structures of CML36, guiding a large structural transition from a molten globule apo-state to a compact holoconformation. Importantly, through binding and activity assays, we showed that CML36 interacts directly with the regulative N terminus of the plasma membrane Ca-ATPase isoform 8 (ACA8) and that this interaction stimulates ACA8 activity. Gene expression analysis revealed that and are co-expressed mainly in inflorescences. Collectively, our results support a role for CML36 as a Ca sensor that binds to and modulates ACA8, uncovering a possible involvement of the CML protein family in the modulation of plant-autoinhibited Ca pumps.