RSL lectin - a complex approach to study carbohydrate-protein interaction

Josef Houser^1,2, Jiří Nováček^1,2, Jan Komárek^1,2, David Buchta³, Dan Pokorný³, Peter Bystrický², Lukáš Žídek^1,2, Michaela Wimmerová^1,2,3

¹CEITEC-Central European Institute of Technology, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic, ²National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Kotlářská 2, 611 37 Brno, Czech Republic, ³Department of Biochemistry, Faculty of Science, Masaryk University, Kotlářská 2, 611 37 Brno, Czech Republic

houser@mail.muni.cz

Lectins, carbohydrate-binding proteins, play an important role in various processes like cell-cell communication or host-pathogen interaction. Due to their specificity and reversibility in binding, they are also widely used for detection, separation and/or analysis of various glycans and glycoproteins [1]. The detailed understanding of interaction between the protein and particular saccharide may help both in analyzing lectin function and lectin engineering for scientific applications.

Fucose-binding lectin AAL from Aleuria aurantia is known for decades and widely used in research [2]. However, its five non-equivalent binding sites make it an uneasy target for detailed analysis of protein-ligand interaction. Its structural homologue RSL form Ralstonia solanacearum, on the other hand, forms only two sets of binding site of very similar amino acid composition [3]. As such, this lectin can be a suitable model for AAL, or maybe even replace it in some applications.

In our study, we focused mainly on RSL ability to bind α- and β-methylfucoside. We created a set of mutants in order to distinguish between both RSL binding sites. Several methods were combined to analyze the interaction of the proteins with saccharides from the thermodynamical and structural point of view. Hence we employed isothermal titration calorimetry (ITC) and nuclear magnetic resonance (NMR) titration as well as X-ray crystallography, where we obtained several high-resolution structures. Combining all these methods, we got a complex view on lectin-carbohydrate interaction in this interesting lectin family.

The research leading to these results obtained financial contribution from the European Union under the Seventh Framework Programme by CEITEC (CZ.1.05/1.1.00/02.0068) project from European Regional Development Fund, SYLICA (Contract No. 286154 under ‘‘Capacities’’ specific programme), Czech Ministry of Education (LH13055) and the Czech Science Foundation (GA13-25401S).

1. H. Lis, N. Sharon. (2007) Lectins, Springer, Netherlands

2. M. Wimmerova, E. Mitchell, J.-F. Sanchez, C. Gautier, A. Imberty, (2003) J. Biol. Chem. 278 (29), 27059–27067

3. N. Kostlánová N, E. P. Mitchell, H. Lortat-Jacob, S. Oscarson, M. Lahmann. N. Gilboa-Garber, G. Chambat, M. Wimmerová, A. Imberty (2005) J. Biol. Chem. 280: 27839-27849