Target: Alpha Synuclein A90C
Nature: Recombinant
Swiss-Prot: P37840-1 (wildtype)
Expression System: E. coli
Protein Length: Full length (1 - 140 aa)
Amino Acid Sequence: MDVFMKGLSKAKEGVVAAAEKTKQGVAEAAGKTKEGVLYVGSKTKEGVVHGVATVAEKTKEQVTNVGGAVVTGVTAVAQKTVEGAGSIACATGFVKKDQLGKNEEGAPQEGILEDMPVDPDNEAYEMPSEEGYQDYEPEA
Purification: Ion-exchange Purified
Purity: >95%
Storage Buffer: 20mM Hepes pH 7.4, 150mM NaCl, 1mM TCEP pH 7.0
Protein Size: 14.49 kDa
Conjugate: No Tag
Scientific Background: Thioflavin-T (ThT) fluorescence remains a common measurement of alpha-synuclein fibril formation, yet ThT exhibits poor affinity for oligomers and early aggregates. The alpha-synuclein A90C mutant monomers can be specifically labelled with alternative fluorophores (such as Alexa 488/Alexa 647) via maleimide chemistry to enable more sensitive FRET analysis of aggregation. The A90C mutant showed no perturbation of monomer structure and Alexa Fluor dye attachment to cysteine 90 was demonstrated to have no effect on the kinetics of fibril formation (1-3). Residue 90 is at the periphery of the NAC region, a key constituent of the alpha-synuclein β-sheet fibril core, which results in fluorophores on different monomers coming into close proximity upon formation of β-sheet structure during aggregation (4).Note - to prevent the potential mis-translation of alpha-synuclein Y136 as C136 during E.coli expression, the Y136-TAT construct was used (5).
References: 1.Thirunavukkuarasu, et al. 2008. Multiparametric Flurorescence Detection of Early Stages in the Amylord Protein Aggregation of Pyrene-labeled α-Synuclein. J. Mol. Biol. 378(5): 1064-73. https://doi.org/10.1016/j.jmb.2008.03.0342.Cremades, et al. 2012. Direct Observation of the Interconversion of Normal and Toxic Forms of α-Synuclein. Cell. 149(5): 1048-59. doi: 10.1016/j.cell.2012.03.0373.Horrocks et al. 2015. Fast Flow Microfluidics and Single-Molecule Fluorescence for the Rapid Characterization of α-Synuclein Oligomers. Anal. Chem. 87(17): 8818-26. https://doi.org/10.1021/acs.analchem.5b018114.Iljina et al. 2016. Kinetic model of the aggregation of alpha-synuclein provides insights into prion-like spreading. PNAS. 113(19): E1206-15. doi: 10.1073/pnas.15241281135.Masuda, et al. 2006. Cysteine misincorporation in bacterially expressed human alpha-synuclein. FEBS Lett. 580(7): 1775-9. doi: 10.1016/j.febslet.2006.02.032
Field of Use: Not for use in humans. Not for use in diagnostics or therapeutics. For research use only.
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