Substrate specificity of glutaminyl cyclases from plants and animals

Glutaminyl cyclases (QC) catalyze the intramolecular cyclization of Nterminal glutamine residues of peptides and proteins. For a comparison of the substrate specificity of human and papaya QC enzymes, a novel continuous assay was established by adapting an existing discontinuous method. Specificity constants (k(cat)/K-m) of dipeptides and dipeptide surrogates were higher for plant QC, whereas the selectivity for oligopeptides was similar for both enzymes. However, only the specificity constants of mammalian QC were dependent on size and composition of the substrates. Specificity constants of both enzymes were equally pHdependent in the acidic pHregion, revealing a pKa value identical to the pKa of the substrate, suggesting similarities in the substrate conversion mode. Accordingly, both QCs converted the Lbetahomoglutaminyl residue in the peptide HbetahomoGlnPheLys ArgLeuAlaNH(2) and the glutaminyl residues of the branched peptide HGlnLys(Gln)ArgLeuAlaNH(2) as well as the partially cyclized peptide HGlncyclo(Nepsilon-LysArgProAlaGlyPhe). In contrast, only QC from C. papaya was able to cyclize a methylated glutamine residue, while this compound did not even inhibit human QC-catalysis, suggesting distinct substrate recognition pattern. The conversion of the potential physiological substrates [Gln(1)]gastrin, [Gln(1)]neurotensin and [Gln(1)]fertilization promoting peptide indicates that human QC may play a key role in posttranslational modification of most if not all pGlucontaining hormones.