Rapid kinetics of calcium dissociation from plant calmodulin and calmodulin-like proteins and effect of target peptides

Abstract

Calcium (Ca²⁺) signaling represents a universal information code in plants, playing crucial roles spanning developmental processes to stress responses. Ca²⁺ signals are decoded into defined plant adaptive responses by different Ca²⁺ sensing proteins, including calmodulin (CaM) and calmodulin-like (CML) proteins. Although major advances have been achieved in describing how these Ca²⁺ decoding proteins interact and regulate downstream target effectors, the molecular details of these processes remain largely unknown. Herein, the kinetics of Ca²⁺ dissociation from a conserved CaM and two CML isoforms from A. thaliana has been studied by fluorescence stopped-flow spectroscopy. Kinetic data were obtained for the isolated Ca²⁺-bound proteins as well as for the proteins complexed with different target peptides. Moreover, the lobe specific interactions between the Ca²⁺ sensing proteins and their targets were characterized by using a panel of protein mutants deficient in Ca²⁺ binding at the N-lobe or C-lobe. Results were analyzed and discussed in the context of the Ca²⁺-decoding and Ca²⁺-controlled target binding mechanisms in plants.

Keywords: Arabidopsis thaliana; Calcium; Calmodulin; Calmodulin-like proteins; Melittin; RAD4; Rapid kinetics; SAC3B.