Investigating the molecular basis of tsetse-trypanosome interactions

Date

2014-09-12

Authors

Goomeshi Nobary, Sarah

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Abstract

The parasitic pathogens of genus Trypanosoma cause significant morbidity and mortality worldwide. The most well studied Trypanosoma related diseases are African sleeping sickness (Trypanosoma brucei) and African Animal Trypanosomiasis (Trypanosoma congolense). Despite more than 100 years of research these diseases continue to have a devastating impact on the socioeconomic development of Africa. A major impediment to controlling outbreaks is the lack of an effective vaccine due, in part, to the parasite’s ability to continually alter its protein coat while in the host, which results in effectively evading the host immune system. Recent studies have identified Trypanosoma congolense proteins that are selectively expressed during transmission in the tsetse arthropod vector where the parasite’s protein coat is not constantly recycled. Of these proteins, Congolense Insect Stage Specific Antigen (TcCISSA) and Congolense Epimastigote Specific Protein (TcCESP) were selected for characterization based on cellular localization, expression levels and predicted roles in facilitating transmission by the tsetse fly. The goal of the present study is to understand the crosstalk between T. congolense and its vector, the tsetse fly. Revealing the structure of proteins is a crucial step in determining their functions. In order to gain insight into the molecular basis of structure and function of TcCESP and TcCISSA we took various biophysical and biochemical approaches. TcCISSA was recombinantly produced in E. coli, crystallized and diffraction quality data collected to 2.5 Å resolution. Structure determination, however, has been problematic due to the absence of homologous models and the inability to take advantage of SelMet phasing due to the presence of only a single methionine in the sequence. Structure determination efforts are ongoing using multiple approaches including NMR. In contrast to TcCISSA, the size and complexity of TcCESP required insect cells for efficient recombinant production. While crystallization trials have yet to yield diffraction quality crystals, a combination of homology modeling validated by chemical crosslinking and mass spectrometry, and circular dichroism spectroscopy have yielded intriguing insight into the architecture of CESP. Characterizing the function of these proteins offers the potential for rare insight into the molecular crosstalk between the parasite and vector and may support the development of novel transmission blocking vaccines.

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Keywords

trypanosoma congolense, tsetse, tsetse-trypanosom interactions, CESP, CISSA

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