Reference : Nanobodies as structural probes to investigate the mechanism of fibril formation by t...
Scientific congresses and symposiums : Poster
Life sciences : Biochemistry, biophysics & molecular biology
http://hdl.handle.net/2268/126169
Nanobodies as structural probes to investigate the mechanism of fibril formation by the amyloidogenic variants of human lysozyme
English
Dumont, Janice mailto [Université de Liège - ULg > Département des sciences de la vie > Enzymologie et repliement des protéines >]
pardon, Els [Vrije Universiteit Brussel - VUB > Department Ultrastructure > > >]
Aumont-Nicaise, Magali [Université Paris-Sud 11 > CNRS - IBBMC > > >]
Menzer, Linda [Université de Liège - ULg > Departement des sciences de la vie > Enzymologie et repliement des protéines > Master2 >]
Kumita, Janet [University of Cambridge > Department of Chemistry > > >]
Willet, Nicolas mailto [Université de Liège - ULg > Département de chimie (sciences) > Nano-chimie et systèmes moléculaires >]
Jérôme, Christine mailto [Université de Liège - ULg > Département de chimie (sciences) > Centre d'études et de rech. sur les macromolécules (CERM) >]
Mazzucchelli, Gabriel mailto [Université de Liège - ULg > > Center for Analytical Research and Technology (CART) >]
Buell, Alexandre [University of Cambridge > Department of Chemistry > > >]
Desmadril, Michel [Université Paris-Sud 11 > CNRS-IBBMC > > >]
De Pauw, Edwin mailto [Université de Liège - ULg > Département de chimie (sciences) > GIGA-R : Laboratoire de spectrométrie de masse (L.S.M.) >]
Wyns, Lode [Vrije Universiteit Brussel - VUB > Department Ultrastructure > > >]
Dobson, Christopher [University of Cambridge > Department of chemistry > > >]
Dumoulin, Mireille mailto [Université de Liège - ULg > Département des sciences de la vie > Enzymologie et repliement des protéines >]
Jun-2012
A0
No
No
International
Gordon Research Conferences: Biopolymers
Du 3 juin 2012 au 08 juin 2012
[en] Amyloid fibrils ; Variants of human lysozyme ; Fragments of camelid antibody
[en] Six variants of human lysozyme (single-point mutatants I56T, F57I, W64R, D67H and double mutants F57I/T70N, W112R/T70N) are associated with a hereditary non-neuropathic systemic amyloidosis. These proteins form extracellular amyloid fibrils that deposit in a wide range of tissues and organs such as liver, spleen and kidneys where they cause damages [1]. It was shown that the D67H and I56T mutations cause a loss in stability and more particularly a loss of global cooperativity of protein [1]. Consequently, under physiologically relevant conditions, these variants can transiently populate a partially unfolded state in which the beta-domain and the C-helix are cooperatively unfolded while the rest of the protein remains native like [1]. The formation of intermolecular interactions between the regions that are unfolded in this intermediate state is likely to be a fundamental trigger of the aggregation process that ultimately leads to the formation and deposition of fibrils in tissues. We have also shown that the binding of three variable domain of camelid antibodies (VHHs) - raised against the wild type human lysozyme inhibit in vitro the formation of amyloid fibrils by the lysozyme variants. These three VHHs bind on different regions of lysozyme and act as amyloid fibril inhibitor through different mechanisms [2, 3, and unpublished results].
In the present work, sixteen new VHHs specific of human lysozyme have been generated. Competition experiments have shown that they bind to five non-overlapping epitopes. We have demonstrated that five of these VHHs are able to bind lysozyme in conditions used for amyloid fibril formation, and interestingly two of them recognize two epitopes that are different from those of the three VHHs previously characterized [2, 3, and unpublished results]. The effects of these new VHHs on the properties of lysozyme variants such as stability, cooperativity and aggregation will be discussed.

[1] Dumoulin, M., J.R. Kumita, and C.M. Dobson, Normal and aberrant biological self-assembly: Insights from studies of human lysozyme and its amyloidogenic variants. Acc Chem Res, 2006, 39(9), 603-610.
[2] Dumoulin, M., et al., A camelid antibody fragment inhibits the formation of amyloid fibrils by human lysozyme. Nature, 2003, 424, 783-788.
[3] Chan, P.H., et al., Engineering a camelid antibody fragment that binds to the active site of human lysozyme and inhibits its conversion into amyloid fibrils. Biochemistry, 2008, 47, 11041-11054.
Centre d'Ingénierie des Protéines - CIP
Fonds pour la formation à la Recherche dans l'Industrie et dans l'Agriculture (Communauté française de Belgique) - FRIA ; Université de Liège Patrimoine
Etude des propriétés amyloidogéniques des variants du lysozyme humain à l'aide de fragments d'anticorps de camélidé comme sondes structurales
Researchers
http://hdl.handle.net/2268/126169

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