N-glycosylation modulates enzymatic activity of Trypanosoma congolense trans-sialidase

Abstract

Trypanosomes cause the devastating disease trypanosomiasis, in which the action of trans-sialidase (TS) enzymes harbored on their surface is a key virulence factor. TS are N-glycosylated, but the biological functions of their glycans has remained elusive. In this study, we investigated the influence of N-glycans on the enzymatic activity and structural stability of TconTS1, a recombinant TS from the African parasite Trypanosoma congolense. The enzyme was expressed in CHO Lec1 cells, which produce high-mannose type N-glycans similar to the TS N-glycosylation pattern in vivo. MALDI-TOF MS data revealed that up to eight putative N-glycosylation sites were glycosylated. N-glycan removal via EndoHf treatment of TconTS1 led to a decrease in substrate affinity relative to the untreated enzyme, but apparently has no impact on the conversion rate. No changes in secondary structure elements of hypoglycosylated TconTS1 were observed in circular dichroism experiments. Molecular dynamics simulations provided evidence for interactions between monosaccharide units of the highly flexible N-glycans and some conserved amino acids located at the catalytic site. These interactions led to conformational changes, possibly enhancing substrate accessibility and enzyme-substrate complex stability. The here-observed modulation of catalytic activity via N-glycans represents a so far unknown structure-function relationship potentially inherent in several members of the TS enzyme family.