1WCS

A mutant of Trypanosoma rangeli sialidase displaying trans-sialidase activity

Summary for 1WCS

• Entry DOI: 10.2210/pdb1wcs/pdb
• Related: 1MZ5 1MZ6 1N1S 1N1T 1N1V 1N1Y
• Descriptor: SIALIDASE (1 entity in total)
• Functional Keywords: trans-sialidase, sialidase, trypanosoma cruzi, trypanosoma rangeli, protein engineering, hydrolase
• Biological source: TRYPANOSOMA RANGELI
• Total number of polymer chains: 1
• Total formula weight: 70071.92

Authors

Paris, G.,Ratier, L.,Amaya, M.F.,Nguyen, T.,Alzari, P.M.,Frasch, C. (deposition date: 2004-11-21, release date: 2004-12-15, Last modification date: 2024-10-09)

Primary citation

Paris, G.,Ratier, L.,Amaya, M.F.,Nguyen, T.,Alzari, P.M.,Frasch, C.
A Sialidase Mutant Displaying Trans-Sialidase Activity
J.Mol.Biol., 345:923-, 2005

PubMed Abstract: Trypanosoma cruzi, the agent of Chagas disease, expresses a modified sialidase, the trans-sialidase, which transfers sialic acid from host glycoconjugates to beta-galactose present in parasite mucins. Another American trypanosome, Trypanosoma rangeli, expresses a homologous protein that has sialidase activity but is devoid of transglycosidase activity. Based on the recently determined structures of T.rangeli sialidase (TrSA) and T.cruzi trans-sialidase (TcTS), we have now constructed mutants of TrSA with the aim of studying the relevant residues in transfer activity. Five mutations, Met96-Val, Ala98-Pro, Ser120-Tyr, Gly249-Tyr and Gln284-Pro, were enough to obtain a sialidase mutant (TrSA(5mut)) with trans-sialidase activity; and a sixth mutation increased the activity to about 10% that of wild-type TcTS. The crystal structure of TrSA(5mut) revealed the formation of a trans-sialidase-like binding site for the acceptor galactose, primarily defined by the phenol group of Tyr120 and the indole ring of Trp313, which adopts a new conformation, similar to that in TcTS, induced by the Gln284-Pro mutation. The transition state analogue 2,3-didehydro-2-deoxy-N-acetylneuraminic acid (DANA), which inhibits sialidases but is a poor inhibitor of trans-sialidase, was used to probe the active site conformation of mutant enzymes. The results show that the presence of a sugar acceptor binding-site, the fine-tuning of protein-substrate interactions and the flexibility of crucial active site residues are all important to achieve transglycosidase activity from the TrSA sialidase scaffold.

PubMed: 15588836
DOI: 10.1016/J.JMB.2004.09.031

Experimental method

X-RAY DIFFRACTION (2.8 Å)