Cysteine misincorporation in bacterially expressed human alpha-synuclein.

Authors :: Masuda M
Dohmae N
Nonaka T
Oikawa T
Hisanaga S
Goedert M
Hasegawa M
Source :: FEBS letters [FEBS Lett] 2006 Mar 20; Vol. 580 (7), pp. 1775-9. Date of Electronic Publication: 2006 Feb 24.
Publication Year :: 2006
Abstract: Bacterially expressed human alpha-synuclein (alpha-syn) has been widely used in structural and functional studies. Here we show that approximately 20% of human alpha-syn expressed in Escherichia coli is mistranslated and that a Cys residue is incorporated at position 136 instead of a Tyr. Site-directed mutagenesis of codon 136 (TAC to TAT) resulted in the expression of alpha-syn lacking Cys. Although wild-type (Y136-TAC and Y136-TAT) and mutant (C136-TGC) alpha-syn had similar propensities to assemble into filaments, the levels of dimeric alpha-syn were increased by misincorporation. To avoid potential artefacts, we recommend use of the Y136-TAT construct for the expression of human alpha-syn.