David Snary

Cell surface antigens of Trypanosoma cruzi: Use of monoclonal antibodies to identify and isolate an epimastigote specific glycoprotein

A monoclonal antibody was produced by cell fusion from mice immunized with Trypanosoma cruzi epimastigotes. The antibody was epimastigote specific but not strain specific; the antibody bound to Y, Peru and Tulahuén epimastigotes but did not bind to Y amastigotes or trypomastigotes. The antigen recognised by the monoclonal antibody was a 72 000 molecular weight cell surface glycoprotein which represented only 0.04% of the whole cell protein. Analysis of the glycoprotein purified by antibody affinity chromatography revealed a carbohydrate content of 52% by weight which was composed of glucosamine, mannose, galactose, glucose and three different pentoses: fucose, xylose and ribose. Immunisation with the purified glycoprotein did not protect mice from a lethal infection of T. cruzi.

Comparative compositions of cell surface glycoconjugates isolated from Trypanosoma cruzi epimastigotes

Cell surface glycoconjugates of epimastigotes of Trypanosoma cruzi have been isolated and analyzed to give their amino acid and carbohydrate compositions. Those which have been investigated are a complex of three closely associated glycoproteins, GP24, GP31, GP37, and a lipopeptidophosphoglycan. The GP24-GP31-GP37 complex has an unusual amino acid composition with very low levels of hydrophobic amino acids, it contains 56% (w/w) carbohydrate, with mannose, galactose and glucosamine (presumably N-acetyl) being present in approximately equal quantities. The lipopeptidophosphoglycan also has low levels of hydrophobic amino acids and contains equal levels of mannose and galactose together with lesser amounts of (N-acetyl) glucosamine. The glycoconjugates are contrasted and compared with two other previously characterised cell surface glycoproteins (GP25 and GP72) from T. cruzi.

An antigenic determinant common to both mouse red blood cells and several membrane proteins of the parasitic protozoa Babesia rodhaini

Four membrane proteins of Babesia rodhaini have been identified by monoclonal antibodies. These proteins are differentiated by serological criteria and by tryptic peptide maps but contain common antigenic determinants. In vitro translation of parasite polyadenylated RNA followed by immunoprecipitation demonstrates that these common determinants are located within the primary sequence of the protein and do not result from post-translational modification. Part of the common amino acid sequence of these proteins is also present within a protein found in mouse red blood cells. Such common structures could potentially play an important role in the host-parasite relationship. The presence of sequence homology within a group of proteins from Babesia rodhaini could be further evidence for the atypical genetic rearrangements proposed to take place within Babesia species.

Formation of mucous gel from the high molecular weight mucoprotein of gastric mucus

The highly viscous and gelatinous nature of gastric mucus is shown to depend upon the properties of a mucoprotein of protein content 13.6 % and molecular weight 2 × 106. At 25°C, the viscosity of this mucoprotein is strongly concentration dependent and at concentrations about 4 g l.–1 rises asymptotically, indicating the onset of a reversible sol-gel transition. Reducing agents (mercaptoethanol) and trypsin split the mucoprotein into four and two sub-units of molecular weight 5.2 × 105 and 9.8 × 105 respectively. These reductions in molecular size are accompanied by loss of the viscous and gelatinous properties. The viscosity of the mucoprotein undergoes two sharp changes in KCl, one between 0 and 0.05 M KCl which is attributed to a polyelectrolyte charge shielding phenomenon. The second transition near 1.0 M KCl is interpreted as a marked increase in asymmetry accompanied by contraction and reduced solvation. Increasing concentrations of CsCl reduce the viscosity of the mucoprotein, bringing about a transition which is essentially complete by 0.1 M CsCl and is as great in extent as both transitions in KCl. In CsCl the highly expanded mucoprotein is contracted to a molecule having the same symmetry but an even smaller volume and lower solvation than in high KCl concentrations. Both salt-induced transitions are non-reversible. A mechanism is discussed in terms of the effect of the Cs+, K+ and Cl– ions on water structure and water-mucoprotein interactions. Guanidinium chloride causes partial expansion of the CsCl treated material with a limited aggregation of four mucoprotein molecules on its removal. The viscosity at finite concentration increases markedly and reversibly with temperature and at 45°C the threshold concentration for reversible gel formation is reduced to 2 g l.–1. There is no evidence for intramolcular changes. The temperature dependence is characteristic of intermolecular entropy driven interactions and the results are interpreted to show that the mechanism of gel formation in gastric mucus involves micelle junctions.