Structural Studies of the Ca²⁺ Regulatory Domain of Drosophila Na⁺/Ca²⁺ Exchanger CALX

Abstract

CALX, the NCX homolog in Drosophila, involves in light-mediated Ca²⁺ homeostasis in sensory neuronal cells. CALX exhibits a unique negative Ca²⁺ regulatory property mediated by Ca2+ binding at its intracellular regulatory domain. Our structural studies of individual CBD1 or CBD2 domain reveal that CBD1 is the only Ca²⁺ binding domain in CALX. Crystal structures of the entire Ca²⁺ regulatory domain CBD12 from two alternative splicing isoforms, CALX1.1 and CALX1.2, demonstrate that CBD1 and CBD2 form an open V-shaped conformation with four Ca²⁺ ions bound on the CBD domain interface. The structures together with Ca²⁺ binding analyses strongly argue that the Ca²⁺ inhibition of CALX is achieved by interdomain conformational change induced by Ca²⁺ binding at CBD1. The conformational difference between the two isoforms also raises a hypothesis that alternative splicing residues adjust the interdomain orientation angle between CBD1 and CBD2 to modify the Ca²⁺ regulatory property of the exchanger. These studies not only establish structural basis to understand the inhibitory Ca²⁺ regulation and the alternative splicing modification of CALX, but also shed light on the general Ca²⁺ regulatory mechanism of other mammalian NCX proteins.