Binding of EGF to its receptor induces dimerization of the normally monomeric receptor. Activation of its intracellular tyrosine kinase then occurs through the formation of an asymmetric kinase dimer in which one subunit, termed the "receiver" kinase, is activated by interaction with the other subunit, termed the "activator" kinase [Zhang, et al. (2006) Cell 125: 1137-1149]. Although there is significant experimental support for this model, the relationship between ligand binding and the mechanics of kinase activation are not known. Here we use luciferase fragment complementation in EGF receptor (EGFR)/ErbB2 heterodimers to probe the mechanics of ErbB kinase activation. Our data support a model in which ligand binding causes the cis-kinase (the EGFR) to adopt the receiver position in the asymmetric dimer and to be activated first. If the EGF receptor is kinase active, this results in the phosphorylation of the trans-kinase (ErbB2). However, if the EGF receptor kinase is kinase dead, the ErbB2 kinase is never activated. Thus, activation of the kinases in the EGFR/ErbB2 asymmetric dimer occurs in a specific sequence and depends on the kinase activity of the EGF receptor.