CONICET | Buscador de Institutos y Recursos Humanos

A novel conformational switch determines Trypanosoma cruzi trans sialidase activity at the covalent intermediate stage Diego J. Alonso de Armiñoa, Darío Estrinb, R. M. S. Alvareza, Adrian E. Roitbergc a. Instituto Superior de Investigaciones Biológicas, INSIBIO (CONICET-UNT). Chacabuco 461, San Miguel de Tucumán, Tucumán, Argentina. b. Instituto de Quimica Fisica de los Materiales, Medioambiente y Energía, INQUIMAE-CONICET, Ciudad Universitaria, Pab 2, C1428EHA, Buenos Aires, Argentina. c. Quantum Theory Project and Departament of Chemistry, University of Florida, Gainesville, Florida, United Status of America. Trypanosoma cruzi is the causative agent of Chagas´ disease. T. cruzi trans-sialidase (TcTS) was shown to be an important factor in the microorganism´s virulence, and has been proposed as a target for drug design, a goal that requires a thorough understanding of the enzyme´s mechanism. Evidence indicates that the catalytic mechanism involves a long-lived covalent intermediate (CI), which is later attacked by an acceptor glycoconjugate, completing the sialic acid transfer. A key question is thus, how does TcTS protect the CI from hydrolysis until the acceptor glycoconjugate can position itself in the active site for the transfer reaction to take place. Previous works hypothesized that water accessibility to the anomeric carbon of sialic acid at the CI stage may be restricted in TcTS compared to TrSA --a structurally and mechanistically very similar enzyme with strict hydrolase activity. Our results show not only that this hypothesis is wrong but suggest a novel switch mechanism sensitive to lactose in CI-TcTS, by which it is deactivated in absence of acceptor ligand and reactivated upon its binding, by virtue of a differential flexibility of the backbone in the open vs closed conformation of CI-TcTS which is not present in CI-TrSA.

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