L-Homocysteine (Hcy) is a non-proteinogenic α-amino acid synthesized from dietary methionine. In healthy humans, high Hcy levels are a risk factor for cardiovascular diseases, stroke and type 2 diabetes. A recent study reports that Hcy reacts with Cys10 of transthyretin (TTR), generating a stable covalent adduct. However, to date the effect of S-homocysteinylation on TTR conformational stability remains unknown. Methods The effect of Hcy on the conformational properties of wt- and L55P-TTR were analysed using a set of biophysical techniques. The cytotoxicity of S-homocysteinylated L55P-TTR was also evaluated in the HL-1 cardiomyocyte cell line, while the effects of the assemblies on kinematic and dynamics properties of cardiac muscle cells were analysed in cardiomyocyte syncytia. Results We found that Hcy stabilizes tetrameric wt-TTR, while it destabilizes the tetrameric structure of the L55P mutant, promoting the accumulation of self-assembly-prone monomeric species. Conclusions Our study demonstrated that S-homocysteinylation of the L55P-TTR mutant impairs protein stability, favouring the appearance of toxic monomers. Interestingly, S-homocysteinylation affected only mutant, not wt-TTR. Moreover, we also show that assemblies of S-homocysteinylated L55P-TTR impair cardiomyocytes functional parameters.

S-Homocysteinylation effects on transthyretin: worsening of cardiomyopathy onset / Manuela Leri, Paola Rebuzzini, Anna Caselli, Simone Luti, Antonino Natalello, Sofia Giorgetti, Loredana Marchese, Silvia Garagna, Massimo Stefani, Paolo Paoli, Monica Bucciantini. - In: BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS. - ISSN 0304-4165. - ELETTRONICO. - 1864(2020), pp. 1-12. [10.1016/j.bbagen.2019.129453]

S-Homocysteinylation effects on transthyretin: worsening of cardiomyopathy onset

Manuela Leri
Investigation
;
Anna Caselli
Investigation
;
Simone Luti
Investigation
;
Massimo Stefani
Writing – Review & Editing
;
Paolo Paoli
Writing – Original Draft Preparation
;
Monica Bucciantini
Writing – Original Draft Preparation
2020

Abstract

L-Homocysteine (Hcy) is a non-proteinogenic α-amino acid synthesized from dietary methionine. In healthy humans, high Hcy levels are a risk factor for cardiovascular diseases, stroke and type 2 diabetes. A recent study reports that Hcy reacts with Cys10 of transthyretin (TTR), generating a stable covalent adduct. However, to date the effect of S-homocysteinylation on TTR conformational stability remains unknown. Methods The effect of Hcy on the conformational properties of wt- and L55P-TTR were analysed using a set of biophysical techniques. The cytotoxicity of S-homocysteinylated L55P-TTR was also evaluated in the HL-1 cardiomyocyte cell line, while the effects of the assemblies on kinematic and dynamics properties of cardiac muscle cells were analysed in cardiomyocyte syncytia. Results We found that Hcy stabilizes tetrameric wt-TTR, while it destabilizes the tetrameric structure of the L55P mutant, promoting the accumulation of self-assembly-prone monomeric species. Conclusions Our study demonstrated that S-homocysteinylation of the L55P-TTR mutant impairs protein stability, favouring the appearance of toxic monomers. Interestingly, S-homocysteinylation affected only mutant, not wt-TTR. Moreover, we also show that assemblies of S-homocysteinylated L55P-TTR impair cardiomyocytes functional parameters.
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Manuela Leri, Paola Rebuzzini, Anna Caselli, Simone Luti, Antonino Natalello, Sofia Giorgetti, Loredana Marchese, Silvia Garagna, Massimo Stefani, Paolo Paoli, Monica Bucciantini
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2158/1176927
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