Glutamine cyclases catalyse the conversion of L-glutaminyl-peptides into 5-oxoprolyl-peptides with the concomitant liberation of ammonia. We report here biophysical characterisation of the glutamine cyclase present in the laticiferous cells of the plant Carica papaya. After purification to near homogeneity, this enzyme was subjected to limited proteolysis and found to exhibit a high resistance to degradation and nicking. The structural reasons for this property were examined using circular dichroism and infrared spectroscopies. By combining the analyses of the infrared and CD spectra of papaya glutamine cyclase, its susceptibility to proteolysis, and its hydrogen-deuterium exchange characteristics, we conclude that this protein contains extensive β-sheet structure and is likely to have only short immobile loops connecting its β-strands.Keywords : secondary structure ; circular dichroism; infrared spectroscopy ; limited proteolysis; hydrogendeuterium exchange.The aerial parts of the tropical plant Carica papaya contain laticiferous cells which constitute a particularly rich source of cysteine proteinases. The well-documented proteinases papain, glycyl endopeptidase, chymopapain and caricain are all secreted by these cells [1Ϫ4]. Besides these proteinases, minor protein constituents including two chitinases [5,6], a glutamine cyclase [7] and a serine proteinase inhibitor [8] have been isolated and partially characterised.Although they are synthesised as inactive preproenzymes, the four papaya proteinases exist in their mature forms in the laticifers [9] because freshly collected papaya latex exhibits full proteolytic activity. The catalytic competence of papain-like cysteine proteinases requires the generation of a catalytic-site imidazolium-thiolate ion pair and the protonic dissociation of a carboxylic acid function with a pKa value close to 4 [10], which controls the ion pair geometry. Three elements help sustain full proteolytic activity in the papaya laticiferous cells: (a) the absence of cysteine proteinase inhibitors, (b) the presence of a high concentration (about 20 mM) of low molecular-mass thiol-containing compounds, and (c) a buffering capacity that maintains the pH around 5.2. From a physiological point of view, papaya laticifers may therefore be regarded as an extreme milieu. The structural reasons for the ability of the latex enzymes to elude -A-benzoyl-DL-arginine-p-nitroanilide; BTPNA, N-A-benzoyl-DL-tyrosine-p-nitroanilide. Enzymes. Glutamine cyclases (EC 2.3.2.5); papain (EC 3.4.22.2) ; glycyl endopeptidase (EC 3.4.22.25); chymopapain (EC 3.4.22.6) ; caricain (EC 3.4.22.30). rapid in vivo degradation through autolysis and proteolysis has not previously been examined.Within this context, we present a study of a recently purified glutaminyl-peptide cyclotransferase from the laticifers of C. papaya (referred to here as papaya glutamine cyclase, or PQC). This enzyme catalyses the conversion of L-glutaminyl-peptides into 5-oxoprolyl-peptides with concomitant liberation of ammonia [11], as shown...