Both RalA and RalB interact with the ubiquitous calcium sensor, calmodulin (CaM). New structural and biophysical characterisation of these interactions strongly suggests that, in the native membrane-associated state, only RalA can be extracted from the membrane by CaM and this non-canonical interaction could underpin the divergent signalling roles of these closely related GTPases. The isoform specificity for RalA exhibited by CaM is hypothesised to contribute to the disparate signalling roles of RalA and RalB in mitochondrial dynamics. This would lead to CaM shuttling RalA to the mitochondrial membrane but leaving RalB localisation unperturbed, and in doing so triggering mitochondrial fission pathways rather than mitophagy.