Endopeptidase variations among different life-cycle stages of African trypanosomes

Lysates of different life-cycle stages of Trypanosoma congolense, Trypanosoma vivax and Trypanosoma brucei were analysed for endopeptidase activity, using reaction conditions which permitted a distinction to be made between lysosomal and non-lysosomal activity [Lonsdale-Eccles, J. D. & Grab, D. J. (1987) Eur. J. Biochem. 169, 467-475]. Hydrolysis of Z-Arg-Arg-NHMec (Z = benzyloxycarbonyl, NHMec = 7-amino-4-methylcoumaryl) and Z-Gly-Gly-Arg-NHMec occurred predominantly at alkaline pH and was observed in lysates of both insect and mammalian infective forms of T. brucei and T. congolense. Compared to their other life-cycle stages, procyclic forms of T. brucei and epimastigote forms of T. congolense exhibited enhanced hydrolysis of these substrates. Low levels of hydrolysis of Z-Arg-Arg-NHMec were observed in the bloodstream and epimastigote forms of T. vivax. The hydrolysis of Z-Gly-Gly-Arg-NHMec in each of the life-cycle stages of T. vivax was generally below detectable levels. In lysates of T. congolense, proteolytic and Z-Phe-Arg-NHMec-hydrolytic activity in bloodstream forms greater than metacyclic greater than epimastigote greater than procyclic forms. In T. vivax Z-Phe-Arg-NHMec-hydrolytic activity differed slightly according to the origin of the parasite but, in general, followed the same pattern (i.e. bloodstream forms greater than epimastigote forms, with metacyclic forms usually intermediate between these two). In T. brucei, Z-Phe-Arg-NHMec-hydrolytic activity in bloodstream forms greater than procyclic forms. Upon differentiation of the long, slender bloodstream forms into short, stumpy forms the Z-Phe-Arg-NHMec-hydrolytic activity was elevated even further. Thus, during their life cycle, each of these African trypanosomes exhibits complex changes of endopeptidase activity, suggestive of an induction of lysosomal activity between the insect and mammalian forms.