Human brain pyroglutamyl peptidase (PAPI; EC 3.4.19.3) is an omega exopeptidase which cleaves pyroglutamic acid from the N-terminus of bioactive peptides and proteins. It plays an important role in the processing and degradation of regulatory peptides such as thyrotropin releasing hormone (TRH) and luteinizing hormone releasing hormone (LHRH). To gain further insights into its performance in vivo and suggest possible applications, such as peptide processing or sequencing, this study focuses on the in vitro stability properties and Michaelis- Menten kinetics of the recombinant wild type enzyme and a single-site mutant, Tyr147→Phe (Y147F).