Binding specificity of mannose-specific carbohydrate-binding protein from the cell surface of Trypanosoma cruzi.
The sugar binding specificity of the recently described mannose-specific carbohydrate-binding proteins (CBP) isolated to homogeneity from both the epimastigote and trypomastigote stages of the pathogenic protozoa Trypanosoma cruzi has been studied by quantitative hapten inhibition of the biotinylated CBPs to immobilized thyroglobulin using model oligosaccharides. The results clearly show a differential specificity toward high-mannose glycans between the CBPs from the two developmental stages. Thus, the isolated CBP from epimastigotes exhibited stronger affinity for higher mannose oligomers containing the Manalpha1-2Manalpha1-6Manalpha1-6 structure. Its affinity decreased, as did the number of mannose residues on the oligomer or removal of the terminal Manalpha1-2-linked mannose. By contrast the CBP isolated from the trypomastigote stage showed about 400-fold lower avidity than the epimastigote form, and contrary to it, it was slightly more specific toward Man5GlcNAc than Man9GlcNAc. Analysis of the interaction of epimastigote-Man-CBP with its ligands by UV difference spectroscopy indicates the existence of an extended binding site in that protein with a large enthalpic contribution to the binding. The thermodynamic parameters of binding were obtained by isothermal titration calorimetry and been found that the DeltaH values to be in good agreement with the van't Hoff values. The binding reactions are mainly enthalpically driven and exhibit enthalpy-enthropy compensation. In addition, analysis of the high-mannose glycans from different parts of the digestive tract of the reduviid insect vector of T. cruzi suggest a role of the CBP in the retention of the epimastigote stage in the anterior portion of the gut.
Bonay
P
P
Centro de Biologia Molecular Severo Ochoa, Universidad Autonoma de Madrid, Madrid, Spain.
Molina
R
R
Fresno
M
M
D000818:Animals; D002151:Calorimetry; D002240:Carbohydrate Sequence; D002352:Carrier Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D008358:Mannose / Q000378:metabolism*; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000378:metabolism; D011485:Protein Binding; D011956:Receptors, Cell Surface; D013056:Spectrophotometry, Ultraviolet; D013954:Thyroglobulin / Q000378:metabolism; D014349:Trypanosoma cruzi / Q000378:metabolism*
D002352:Carrier Proteins; D011134:Polysaccharides; D011956:Receptors, Cell Surface; C055213:saccharide-binding proteins; D013954:Thyroglobulin; D008358:Mannose