Reference : Production of four amyloidogenic variants of human lysozyme as inclusion bodies in Esche...
Scientific congresses and symposiums : Poster
Life sciences : Biochemistry, biophysics & molecular biology
http://hdl.handle.net/2268/104042
Production of four amyloidogenic variants of human lysozyme as inclusion bodies in Escherichia coli
English
Dumont, Janice mailto [Université de Liège - ULg > Département des sciences de la vie > Enzymologie et repliement des protéines >]
Menzer, Linda mailto [Université de Liège - ULg > > > 2e an. master bioch. & biol. moléc. & cell., fin. appr.]
Scarafone, Natacha mailto [Université de Liège - ULg > > > Doct. sc. (bioch., biol. mol.&cell., bioinf.&mod.-Bologne)]
Duez, Colette mailto [Université de Liège - ULg > > Centre d'ingénierie des protéines >]
Dumoulin, Mireille mailto [Université de Liège - ULg > Département des sciences de la vie > Enzymologie et repliement des protéines >]
2009
No
International
Penicillin-recognizing enzymes: from enzyme kinetics to protein folding.
1-3 juillet 2009
Université de Liège
Liège
Belgique
[en] Amyloid Fibril ; Nanobodies ; Lysozyme
[en] Six variants of human lysozyme (I56T, F57I, W64R, D67H, F57I/T70N and W112R/T70N) are associated with a hereditary non-neuropathic systemic amyloidosis. This disease involved an extra cellular deposition of amyloid fibrils made of lysozyme variants in a wide range of organs such as liver, spleen and kidneys [1]. The characterisation at the molecular level of two variants, I56T and D67H, has shown that these mutations reduce the stability and more particularly the global cooperativity of the protein. 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.
In order to study the effects of the other amyloidogenic mutations on the properties of lysozyme and thus to get more insight in the mechanism of amyloid formation, it is necessary to produce them in large quantities. The D67H, I56T and F57I variants are currently produced in Aspergillus niger; the expression in this organism is, however, time consuming and the yield is very low. The attempts to use alternative systems such as Pichia pastoris [2], Saccharomyces cerevisiae, and Arabidopsis thaliana have not been conclusive so far.
In this work, we have produced the four single-point lysozyme variants as inclusion bodies in Escherichia coli and explored the possibility to refold them.
[1] Dumoulin & al., (2006) Acc. Chem. Res., 39, 603 - 610
[2] Kumita & al., (2006) FEBS J., 273, 711-720
http://hdl.handle.net/2268/104042

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