Dimerization of Tyr136Cys alpha-synuclein prevents amyloid transformation of wild type alpha-synuclein

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• 作者：K.V. Barinova^a ; ^b ; M.L. Kuravsky^a ; A.M. Arutyunyan^a ; M.V. Serebryakova^a ; E.V. Schmalhausen^a ; V.I. Muronetz^a ; ^b ; ^{vimuronets@belozersky.msu.ru}
• 关键词：DTT ; dithiothreitol ; DTNB ; 5 ; 5&prime ; -dithiobis-2-nitrobenzoic acid ; PBS ; phosphate buffered saline (10  ; mM potassium phosphate ; 0.137  ; M NaCl ; 2.7  ; mM KCl) ; Tyr136-AS and Cys136-AS ; recombinant human alpha-synuclein forms containing tyrosine or cysteine residues in position 136 respectively ; ThT ; thioflavin T
• 刊名：International Journal of Biological Macromolecules
• 出版年：2017
• 出版时间：March 2017
• 年：2017
• 卷：96
• 期：Complete
• 页码：35-43
• 全文大小：1072 K
• 卷排序：96

文摘

Expression of human alpha-synuclein in E. coli cells is known to result in a mixture of the wild type alpha-synuclein and the protein containing Tyr136Cys substitution due to the translational error. The amount of Cys136 alpha-synuclein (Cys136-AS) may reach approximately 50% of the recombinant protein. The wild-type and Cys136-containing fractions of alpha-synuclein were separated using thiol-Sepharose, and their properties were investigated. In the absence of reducing agents, Cys136-AS forms dimers due to the disulfide bonding. Both wild-type and Cys136 alpha-synuclein preparations are prone to aggregate during prolonged incubation under shaking at pH 4 and 37 °C, but only the wild-type alpha-synuclein produces amyloid aggregates. The aggregates produced by either monomeric or dimeric Cys136-AS do not exhibit amyloid properties according to the test with Thioflavin T. Moreover, an admixture of dimeric Cys136-AS prevents the amyloid transformation of the wild-type alpha-synuclein. CD spectroscopy analysis revealed an enhanced content of alpha-helical structures in the aggregates produced by dimeric Cys136-AS. The admixture of Cys136-AS in preparations of human recombinant alpha-synuclein can be a source of erroneous interpretation of experiments on amyloid transformation of this protein.