821.1 - Serial crystallography and kinetics reveal how the FimH bacterial lectin tweaks between mono- and multivalent binding of high-mannose N-glycans
Background : The lectin domain of the fimbrial adhesin FimH from Escherichia coli recognizes with the highest affinity and in seemingly monovalent fashion oligomannosides-3 and -5 N-glycans. This happens with a high affinity (Kd=20 nM) because the alpha1,3-linked arm remains free from any further alpha1,2-mannose substitution. The latter causes an at least 10-fold affinity loss in creating the oligomannose-6 N-glycan (1).
Methods : The binding of FimH to pauci- and oligomannoside-containing N-glycans and similarly glycosylated proteins, as well as its dimannoside endings Manalpha1,2, Manalpha1,3 and Manalpha1,6, have been profiled using the FimH Lectprofile kit (GlycoDiag, Applied Glycomics, France) and a newly designed paucimannose-containing glycan array. The kinetics of binding, ka and kd, were analysed using surface plasmon resonance and compared with earlier incomplete data (2).
Results : Mono- and multivalent binding were found in co-crystal structures of the FimH lectin domain with alpha1,6-fucosylated oligomannose-3 and with oligoannose-6, respectively. The kinetics of binding indicate that once alpha1,2-linked mannose comes into play, the complexes are not so stable, as previously also measured using calorimetry (3), and accompanied with a high dissociation constant. Molecular dynamics simulations further support that it is not the total interaction energy that determines the final retained complex (metastable complexes), but the kinetics of interaction with the three possible dimannoside endings.
Conclusions : Kinetics and stability of the complexes drive the most favoured arrangement, via either a unique or multivalent interaction, between the N-glycan and the FimH protein in the crystal.
References:
(1) Bouckaert, J., Mackenzie, J., de Paz, J. L., Chipwaza, B., Choudhury, D., Zavialov, A., Mannerstedt, K., Anderson, J., Pierard, D., Wyns, L., Seeberger, P. H., Oscarson, S., De Greve, H., Knight, S. D. (2006) The affinity of the FimH fimbrial adhesin is receptor-driven and quasi-independent of Escherichia coli pathotypes. Molecular Microbiology 61, 1556-1568
(2) Sauer, M. M., Jakob, R. P., Luber, T., Canonica, F., Navarra, G., Ernst, B., Maier, T., Glockshuber, R. (2019) Binding of the bacterial adhesin FimH to its natural, multivalent high-mannose type glycan targets. J Am Chem Soc 141, 936-944
(3) Dumych, T., Bridot, C., Gouin, S. G., Paryzhak, S., Szunerits, S., Bilyy, R., Bouckaert, J., (2018) A novel integrated way for deciphering the glycan code for the FimH lectin. Molecules 23, 2794
This work was funded in part through contract namp;amp;[deg] 200445 from the Agence Nationale de la Recherche (ANR), on project amp;ldquo;E-clickamp;rdquo;: Electrochemically promoted tyrosine-click chemistry. The authors acknowledge financial support through the Programme for EArly-stage Researchers in Lille (PEARL), under the auspices of the European Union Horizon 2020 research and innovation programme, project "ATACA" with Marie Sklodowska-Curie agreement No. 847568. PEARL is coordinated by the Foundation I-SITE ULNE and implemented at the Centre National de la Recherche Scientific (CNRS). We gratefully acknowledge use of PROXIMA-1 and -2a beamlines at the SOLEIL synchrotron (Saint-Aubin, France) and of beamlines P13 and P14 at the PETRA III storage ring (DESY, Hamburg, Germany).
