Human CD38 is a multifunctional ectoenzyme responsible for catalyzing the conversions from nicotinamide adenine dinucleotide (NAD) to cyclic ADP-ribose (cADPR) and from cADPR to ADP-ribose (ADPR). Both cADPR and ADPR are calcium messengers that can mobilize intracellular stores and activate influx as well. In this study, we determined three crystal structures of the human CD38 enzymatic domain complexed with cADPR at 1.5-Å resolution, with its analog, cyclic GDP-ribose (cGDPR) (1.68 Å) and with NGD (2.1 Å), a substrate analog of NAD. The results indicate that the binding of cADPR or cGDPR to the active site induces structural rearrangements in the dipeptide Glu 146-Asp147 by as much as 2.7 Å, providing the first direct evidence of a conformational change at the active site during catalysis. In addition, Glu226 is shown to be critical not only in catalysis but also in positioning of cADPR at the catalytic site through strong hydrogen bonding interactions. Structural details obtained from these complexes provide a step-by-step description of the catalytic processes in the synthesis and hydrolysis of cADPR.