title: A new class of mechanism-based inhibitors for Trypanosoma cruzi trans-sialidase and their influence on parasite virulence.
abstract: One of the most interesting aspects of Trypanosoma cruzi is its adaptation to obtain sialic acid from its host, fulfilling this need exclusively through the reaction catalyzed by enzymatically active trans-sialidase (aTS), thought to play an important role in the pathogenesis of Chagas' disease. Herein, we report that 2-difluoromethyl-4-nitrophenyl-3,5-dideoxy-d-glycero-alpha-d-galacto-2-nonulopyranosid acid (NeuNAcFNP) inactivates aTS time- and dose-dependently, and this inhibition was not relieved removing the inhibitor. Also, NeuNAcFNP causes a decrease in infection of mammalian cells. Characterization of labeled aTS by matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry revealed that inactivation of the enzyme occurs through formation of a covalent bond between Arg245 and Asp247 and the inhibitor aglycone. Participation of Asp247 in the catalytic mechanism was proved by constructing a TSD247A mutant, which presents only residual activity. Molecular dynamic simulations indicate that the D247A mutation results in a more open catalytic cleft. In summary, NeuNAcFNP is the first reported mechanism-based inhibitor of aTS, representing a new template for drug design and opening new possibilities for chemotherapy of Chagas' disease, as well as for the elucidation of aTS function in T. cruzi pathogenesis and biology.
authors.affiliation: Instituto de Biofísica Carlos Chagas Filho, Centro de Ciências da Saúde, Bloco G, Rio de Janeiro, RJ, Brasil.
authors.forename: Sebastião T
authors.initials: ST
authors.lastname: Carvalho
authors.forename: Mauro
authors.initials: M
authors.lastname: Sola-Penna
authors.forename: Isadora A
authors.initials: IA
authors.lastname: Oliveira
authors.forename: Samuel
authors.initials: S
authors.lastname: Pita
authors.forename: Arlan S
authors.initials: AS
authors.lastname: Gonçalves
authors.forename: Bianca C
authors.initials: BC
authors.lastname: Neves
authors.forename: Francisco R
authors.initials: FR
authors.lastname: Sousa
authors.forename: Leonardo
authors.initials: L
authors.lastname: Freire-de-Lima
authors.forename: Masaki
authors.initials: M
authors.lastname: Kurogochi
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Hinou
authors.forename: Shin-Ichiro
authors.initials: S
authors.lastname: Nishimura
authors.forename: Lucia
authors.initials: L
authors.lastname: Mendonça-Previato
authors.forename: Jose O
authors.initials: JO
authors.lastname: Previato
authors.forename: Adriane R
authors.initials: AR
authors.lastname: Todeschini
mesh_terms: D000818:Animals; D055162:Biocatalysis / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D006023:Glycoproteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D006790:Host-Parasite Interactions / Q000187:drug effects*; D056004:Molecular Dynamics Simulation; D015394:Molecular Structure; D009439:Neuraminidase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D012794:Sialic Acids / Q000737:chemistry / Q000494:pharmacology*; D013329:Structure-Activity Relationship; D014349:Trypanosoma cruzi / Q000187:drug effects / Q000201:enzymology* / Q000472:pathogenicity*
chemical_list: C554547:2-difluoromethyl-4-nitrophenyl-3,5-dideoxygalacto-2-nonulopyranosid acid; D004791:Enzyme Inhibitors; D006023:Glycoproteins; D012794:Sialic Acids; C069715:trans-sialidase; D009439:Neuraminidase