Substrate specificity profiling of M32 metallocarboxypeptidases from Trypanosoma cruzi and Trypanosoma brucei

Publication date: Available online 12 December 2017 Source:Molecular and Biochemical Parasitology Author(s): Alejandra P. Frasch, León A. Bouvier, Florencia M. Oppenheimer, Maria Aparecida Juliano, Luiz Juliano, Adriana K. Carmona, Juan José Cazzulo, Gabriela T. Niemirowicz Metallocarboxypeptidases (MCPs) of the M32 family, while broadly distributed among prokaryotic organisms, have so far been only found in a few eukaryotes including trypanosomatids. Among these organisms are human and animal pathogens of medical relevance such as Trypanosoma brucei and Trypanosoma cruzi, the respective causative agents of sleeping sickness and Chagas disease. The M32 MCP orthologues found in these parasites share 72% protein sequence identity. They also present a cytosolic localization, a similar pattern of expression and a marked preference for Arg/Lys residues at P1'. To further explore MCPs substrate specificity beyond the S1′ subsite, we employed four positional scanning synthetic combinatorial libraries (PS-SC) of fluorescence resonance energy transfer (FRET) peptides. Our results indicated that the T. brucei enzyme has a restricted selectivity for Phe in P1 position compared to T. cruzi MCP-1, which presented a wider range of substrate acceptance. The S2, S3 and S4 subsites, on the other hand, could accommodate a broad range of residues. On the basis of these results, we synthesized for each enzyme a series of FRET substrates which contained the most favourable residues ...
Source: Molecular and Biochemical Parasitology - Category: Parasitology Source Type: research