Comparative structural analysis of surface coat proteins in African trypanosomes

Abstract text

African trypanosomes are causative agents of Human African Trypanosomiasis (HAT) and Animal African Trypanosomiasis (AAT) in sub-Saharan Africa. The cell surface of these trypanosomes is coated with a thick layer of prototypic, glycosylphosphatidylinositol-anchored proteins known as Variant Surface Glycoprotein (VSG), with a molecular size of ca. 60 kDa. The VSG coat acts to counteract the host immune response through antigenic variation by switching the expression between antigenically distinct VSGs from a repertoire of more than 1200 VSG genes. It also serves as a physical barrier to protect underlying invariant proteins from attacks by the host's immune system. In addition, directional swimming of the trypanosomes causes antibodies bound to the VSG coat to be dragged to the posterior pole of the trypanosome where they are endocytosed. The structures of VSGs in Trypanosoma brucei have been extensively studied, which consist of two or three domains, one N-terminal domain consisting of 350–400 residues, followed by one or two C-terminal domains each having 30–70 residues. The NTDs are divided into five main types (N1-N5) and CTDs are divided into six groups (Types 1-6). The VSGs in T. congolense and T. vivax are somewhat smaller, and are made up of a single domain that is similar to the T. brucei VSG N-terminal domain. Although the tertiary structure of NTDs of VSGs in T. brucei has been extensively studied, we have no structural information on either T. congolense or T. vivax VSGs, therefore, we are aiming to solve the structures of VSGs from these species. The first goal is to solve the structure of a number of different, selected T. congolense VSGs. The next step will focus on determining the structure of a T. vivax VSG. A genetically engineered cell line of T. brucei was constructed to express a T. congolense VSG, which was used to purify the VSG in its soluble form (sVSG). Purification of full-length sVSG was performed by anion exchange chromatography. An X-ray crystallography approach will be used to solve the structures of VSGs. Resolving the structural model of VSGs in T. congolense and T. vivax will provide a better understanding of the overall architecture and structure-function relationship of the surface coat in these species.