This thesis investigates the kinetic and stability characteristics of recombinant human brain pyroglutamyl peptidase PAPI, an omega exopeptidase which cleaves pyroglutamic acid from the N-terminus of peptides and proteins. Three classes of pyroglutamyl peptidase have been found in a variety of bacteria, plant, animal, and human tissues; the first class includes the bacterial and animal type 1, pyroglutamyl peptidase I. The genes encoding bacterial PAPI have been cloned and characterized previously, allowing the study of the primary structure of this enzyme and its over-expression in heterologous organisms. Researchers have also been able to clone and characterize the human form of the peptidase. Recombinant human PAPI was over expressed in E. coli grown in LB culture medium and purified by nickel affinity chromatography. The enzyme has a molecular weight of 23kDa, by SDS-PAGE. The estimated T5o was 60°C and the half-life at this temperature was 15 min, £ = 0.046 ± 0.002 min'1. With regard to solvent tolerance, PAPI was tested in dimethyl sulphoxide, methanol, acetone, tetrahydrofuran, acetonitrile, dimethyl formamide and ethanol over a range of v/v concentrations. It was not stable in most solvents and methanol and DMSO were the least injurious for PAPI activity: 56% and 50% residual PAPI activity remained at 10% v/v methanol and DMSO, respectively. Chemical modification with dimethyl suberimidate gave only 2 0 % recovery of initial activity and did not stabilize the enzyme. The effect of polyol additives was investigated and it was found that the enzyme’s activity and stability increased with xylitol. Trehalose, glycerol and ammonium sulphate did not stabilize PAPI. Michaelis-Menten kinetics of PAPI were determined at pH 8.0 with pyroglutamyl 7- aminomethylcoumarin as substrate, Km= 0.132 ± 0.024 mM and ¿cat= 2.68x1 O'5s’1. Both mutant Phel6 —»Tyr (F16Y) and Tyrl47-»Phe (Y147F) were compared with wild type PAPI. The T50of F16Y was 51°C and the half-life at 70°C was 27 min, k= 0.026 ± 0.002 min'1. For Y147F, T5 0 was 78°C and the half-life at 70°C was 25 min, k = 0.028 ± 0.001 min'1. Kinetic values for F16Y were Km= 0.162 ± 0.020 mM and kcat= 5.75xl0' 5 s'1, while Y147F had Km = 0.115 ± 0.019 mM and &cat= 2.45xl0~5 s'1.