The existence of a fifth essential protein for pantothenate biosynthesis in some enteric bacteria has recently been reported by Stuecker et al. [10] and Nozaki et al. (in press) [9]. This protein, PanZ, catalyses the activation of the PanD zymogen to form ADC and is essential for prototrophic growth. In this paper, we characterise the interaction of PanZ with coenzyme A and a constitutively inactive mutant of PanD using a combination of isothermal titration calorimetry and mass spectrometry. These approaches reveal that the two proteins interact with nanomolar affinity in a CoA-dependent fashion to form a heterooctameric complex. (c) 2012 Elsevier Inc. All rights reserved.