Multivalent sialylation of β-thio-glycoclusters by Trypanosoma cruzi trans sialidase and analysis by high performance anion exchange chromatography.
The synthesis of multivalent sialylated glycoclusters is herein addressed by a chemoenzymatic approach using the trans-sialidase of Trypanosoma cruzi (TcTS). Multivalent β-thio-galactopyranosides and β-thio-lactosides were used as acceptor substrates and 3'-sialyllactose as the sialic acid donor. High performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) was shown to be an excellent technique for the analysis of the reaction products. Different eluting conditions were optimized to allow the simultaneous resolution of the sialylated species, as well as their neutral precursors. The TcTS efficiently transferred sialyl residues to di, tri, tetra and octa β-thiogalactosides. In the case of an octavalent thiolactoside, up to six polysialylated compounds could be resolved. Preparative sialylation reactions were performed using the tetravalent and octavalent acceptor substrates. The main sialylated derivatives could be unequivocally assigned by MALDI mass spectrometry. Inhibition of the transfer to the natural substrate, N-acetyllactosamine, was also studied. The octalactoside caused 82 % inhibition of sialic acid transfer when we used equimolar concentrations of donor, acceptor and inhibitor.
CIHIDECAR-CONICET, Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Pabellón 2, Ciudad Universitaria, 1428, Buenos Aires, Argentina. ragusti@qo.fcen.uba.ar.
Rosalía
R
Agustí
CIHIDECAR-CONICET, Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Pabellón 2, Ciudad Universitaria, 1428, Buenos Aires, Argentina.
María Emilia
ME
Cano
CIHIDECAR-CONICET, Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Pabellón 2, Ciudad Universitaria, 1428, Buenos Aires, Argentina.
Alejandro J
AJ
Cagnoni
Laboratoire de Glycochimie, des Antimicrobiens et des Agroressources (LG2A)-CNRS UMR 7478, Université de Picardie Jules Verne, 33 rue Saint Leu, 80039, Amiens Cedex, France.
José
J
Kovensky
CIHIDECAR-CONICET, Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Pabellón 2, Ciudad Universitaria, 1428, Buenos Aires, Argentina.
Rosa M
RM
de Lederkremer
CIHIDECAR-CONICET, Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Pabellón 2, Ciudad Universitaria, 1428, Buenos Aires, Argentina. mluhrig@qo.fcen.uba.ar.
María Laura
ML
Uhrig
D002851:Chromatography, High Pressure Liquid; D006023:Glycoproteins / Q000737:chemistry*; D007785:Lactose / Q000031:analogs & derivatives* / Q000737:chemistry; D009439:Neuraminidase / Q000737:chemistry*; D015800:Protozoan Proteins / Q000737:chemistry*; D012794:Sialic Acids / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013862:Thiogalactosides / Q000737:chemistry*; D014349:Trypanosoma cruzi / Q000201:enzymology*
D006023:Glycoproteins; D015800:Protozoan Proteins; D012794:Sialic Acids; D013862:Thiogalactosides; C000020:N-acetylneuraminoyllactose; C069715:trans-sialidase; D009439:Neuraminidase; D007785:Lactose
Enzymatic sialylation; HPAEC; MALDI-TOF; Multivalent glycoclusters; Sialic acid; T. cruzi trans-sialidase; β-galactopyranosides