The power is in detail - structural analysis of an Aleuria aurantia lectin mutant

Josef Houser^1,2, Radek Matuška², Stanislav Kozmon^1,2, Patric Romano3, Jaroslav Koča^1,2,Michaela Wimmerová^1,2,4

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,
³Institute for Hepatitis and Virus Research, 3805 Old Easton Road, Doylestown, PA 18902 USA and ⁴Department of Biochemistry, Faculty of Science, Masaryk University, Kotlářská 2, 611 37 Brno, Czech Republic

houser@mail.muni.cz

Lectins are able to specifically and reversibly bind carbohydrates and therefore are widely used for detecting, labelling and isolation of glycoproteins and other sugar containing moieties. Changes in glycosylation are connected to various processes including cancer development. Among sugar moieties exhibited on the cell surface, the important role is played by L-fucose (6-deoxy-L-galactose) [1]. Most known fucose-specific lectin is AAL isolated from Aleuria aurantia. Despite the long-term investigation of this protein including the solution of its structure [2], more detailed characterization is demanded in order to create varieties of more strict specificity and/or increased affinity.

The preparation of several artificial constructs based on AAL protein was reported [3]. We have crystallized one such mutant that has been proved to have stronger affinity towards α1-6 bound fucose. We were able to collect several datasets of high quality with resolution up to 1.09Å. The detailed insight into the structure revealed the principal of the affinity change, shading the light upon variability of AAL’s binding sites differences. Furthermore, we utilized in silico approach to describe the contribution of particular amino acids in each of five AAL's binding sites. The combination of experimental and computational techniques is a powerful tool to decipher a complex interaction, where both hydrogen bonds and dispersion forces take part. This research helps us to improve our knowledge of not yet fully understood background of sugar binding variability among AAL lectin family which includes also proteins from opportunistic pathogens such as Burkholderia ambifaria [4] or Aspergillus fumigatus.

This work was supported by the European Community's Seventh Framework Program under the "Capacities" specific programme (Contract No. 286154) and CEITEC - Central European Institute of Technology with research infrastructure supported by the project CZ.1.05/1.1.00/02.0068 from European Regional Development Fund. The project is supported within the SoMoPro programme (project No. 2SGA2747, co-funded by FP7/2007-2013 under grant agreement No. 229603). The authors thank the Czech National Supercomputing Centre, METACENTRUM, for providing computational resources (research intent LM2010005).

1. D. J. Becker, J. B. Lowe, (2003) Glycobiology 13 (7), 41R–53R

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

3. P. Romano, A. Mackay, M. Vong, J. DeSa, A. Lamontagne, M. A. Comunale, J. Hafner, T. Block, R. Lec, A. Mehta. (2011) Biochem. Biophys. Res. Commun. 414 (1), 84–9

4. A. Audfray, J. Claudinon, S. Abounit, N. Ruvoen-Clouet, G. Larson, D. F. Smith, M. Wimmerova, J. Le Pendu, W. Romer, A. Varrot, A. Imberty, (2012) J. Biol. Chem. 287, 4335–4347