Absence of an essential thiol in human glutaminyl cyclase: implications for mechanism

Recent kinetic studies of the papaya glutaminyl cyclase suggest a catalytic mechanism consisting of a direct cyclization of glutaminyl to pyroglutamyl residues, a suggestion consistent with a lack of catalytic nucleophiles at the active site of the plant enzyme. The mammalian glutaminyl cyclase has been reported to possess reactive thiols (Busby et al (1987) J.BioLChem. 262, 85328536) which may imply a different catalytic mechanism from that of the plant enzyme. We have partially sequenced several mammalian and one avian species and found that the two cysteine residues of the human glutaminyl cyclase are completely conserved. However, mutagenesis of these cysteine residues results in only a slight decrease in enzyme activity. Likewise, the recombinant human enzyme was completely resistant to attempted chemical modification of the putative reactive thiols, suggesting the animal and plant glutaminyl cyclases may indeed have similar catalytic mechanisms. Further evidence for this suggestion was the rinding of a nearly identical deuterium solvent isotope effect (1.2 vs 1.3) upon the reaction by the human and papaya enzymes, respectively. Although the human glutaminyl cyclase did not appear to have reactive thiols, it was sensitive to diethylpyrocarbonate and acetylimidazole, suggesting functionally important histidine and tyrosine residues, respectively.