This report describes the characterization of a member of the alpha-crystallin small heat shock protein family in a trypanosomatid, which was isolated from the human pathogen Trypanosoma cruzi. One alpha-crystallin small heat shock protein gene was identified in a database search. The coding region is located in an open reading frame of 429bp encoding a protein of 142 amino acids. The amino acid sequence was deduced from the isolated gene. The protein has an alpha-crystallin domain characteristic of the alpha-crystallin small heat shock proteins and a molecular weight of 15.9kDa, so the protein was designated SHSP16. Analysis of the nucleotide sequences of four different T. cruzi strains showed two different sequences, which correspond to the two main T. cruzi genetic groups. Gene expression analysis by RT-PCR showed increased transcription of the gene after the parasite was exposed to heat stress. Recombinant SHSP16 showed molecular chaperone activity in vitro, because it inhibited the thermal aggregation of the mitochondrial malate dehydrogenase enzyme.