Escherichia coli (E. coli) prephenate dehydrogenase (TyrA) is a dimeric enzyme that catalyzes the oxidative decarboxylation of prephenate to 4-hydroxyphenylpyruvate using NAD+ as a cofactor. TyrA and chorismate mutase (AroQT) form a single-polypeptide bi-functional enzyme known as T-protein. Although experimental and theoretical evidence indicates that the E. coli AroQT and TyrA domains are interdependent, we recently described the successful replacement of the AroQT domain of the T-protein with the small protein-G ?/? immunoglobulin-binding ?1 domain (G?1). The G?1wt-TyrA fusion protein folded into a dimeric conformation and showed such strong TyrA activity that we were able to discard important roles for any interface contact formed between AroQT and TyrA. We also found that the replacement of AroQT with EF-hand Ca2+-binding motifs produce a dimeric, functional E. coli TyrA protein. Interestingly, we found a calcium-dependent switch-off of TyrA activity in constructs containing a canonical-EF-hand motif fused to TyrA. Currently, we are using the TyrA reporter system to study the coiled-coil amino acid sequence space that determines homo-dimer specificity instead of alternative conformations.