Identification and characterization of two repetitive non-variable antigens from African trypanosomes which are recognized early during infection

Abstract

SUMMARYThe present paper describes two repetitive proteins representing common antigens of African trypanosomes which are non-variant and which are recognized early in infection by the host immune system. These antigens were identified by their ability to immunoreact with bovine serum taken during the early phase of a cyclic trypanosomal infection. Screening of a cDNA library from T. b. gambiense with such early infection serum identified a protein which contains a repetitive motif consisting of 68 amino acid repeat units (GM6). Immunofluorescence and immunogold electron microscopy revealed that GM6 is located on fibres which connect the microtubules of the membrane skeleton with the flagellum. A second repetitive antigen detected by this serum is MARP1 (microtubule-associated repetitive protein 1), a protein previously characterized in this laboratory as a high-molecular weight component of the membrane skeleton, which consists of more than 50 tandemly repeated, near-identical 38 amino acid repeat units. Beta-galactosidase fusion products of both proteins demonstrated a strong immunoreactivity with sera from T. b. brucei and T. congolense-infected cattle. The result from this preliminary immunological evaluation indicates a high immunodiagnostic sensititivy (90%) of the two recombinant antigens which make them interesting candidates for immunodiagnosis of trypanosomiasis in cattle.

Evolutionary Relationships and Protein Domain Architecture in an Expanded Calpain Superfamily in Kinetoplastid Parasites

“…Secondary structure analysis predicted a high probability of the repetitive arrays to form a coiled-coil structure. Database searches revealed that short fragments containing some of the repeats of TbCALP11.2, LmCALP27.2, and TcCLP441.10 had been published as antigens GM6 in T. brucei, Lcr1 in Leishmania chagasi, and FRA in T. cruzi, respectively (Lafaille et al 1989;Muller et al 1992;Wilson et al 1995). It was shown in these studies that GM6 is associated with the cellular microtubule structures and FRA with the flagellar cytoskeleton.…”

“…Two of these sequences, TbCALP11.2 and TcCALP441.10, have previously been characterized as cytoskeleton-associated proteins GM6 and FRA, respectively (Lafaille et al 1989;Muller et al 1992). At the time of these publications, only fragments of the repeat were identified by cDNA sequencing and their calpainlike properties were not recognized.…”

“…Despite their overall similarity in sequence and domain structure, the localization of both proteins is different. GM6 is located on fibers which connect the microtubules of the membrane skeleton with the flagellum, whereas FRA (flagellar repetitive antigen) is localized inside the flagellum (Lafaille et al 1989;Muller et al 1992). The presence of repeats in the other five calpain-like proteins of this group, but also the observation that a number of cytoskeleton-associated proteins in trypanosomatids have an internal repeat structure (Gull 1999), suggests that these repeats mediate interaction with microtubules and that the presence of calpain domain II adds an additional element of functionality to these proteins.…”

Improved serodiagnosis of alveolar echinococcosis of humans using anin vitro-producedEchinococcus multilocularisantigen

“…The affinity purification of specific antibodies against the 2 immunodominant bands in the EmVF antigen was performed essentially as described by Mü ller et al (1992). Briefly, the region corresponding to the 20-22 and 8 kDa antigens was excised from the nitrocellulose, following sodium dodecyl-sulfate polyacrylamide gel electrophresis (SDS-PAGE) and blotting of the EmVF antigen.…”

Cell fractionation of parasitic protozoa: a review

“…Furthermore, it is evident that such an intricate structure must have minor components. For T. brucei, those minor components were suggested to be spectrin-type proteins (Schneider et al 1988, Woods et al 1989, Müller et al 1992, Woodward et al 1994, Imboden et al 1995.…”