Synfacts 2020; 16(09): 1129
DOI: 10.1055/s-0040-1706794
Flow Chemistry

Protein Synthesis on Demand, and Make it Snappy!

Contributor(s):
Dirk Trauner
,
Bryan S. Matsuura
Pentelute BL. * et al. Massachusetts Institute of Technology, Cambridge, USA
Synthesis of Proteins by Automated Flow Chemistry.

Science 2020;
368: 980-987
 

Significance

Solid-phase peptide synthesis (SPPS) is routinely performed for the preparation of small peptides. Despite decades of optimization, synthesis of peptides with sequences longer than 50 amino acids is challenging due to the generation of byproducts stemming from deletion, epimerization, and truncation. The chemical synthesis of full-length proteins did not become a practical reality until the introduction of native chemical ligation (NCL), which often requires conditions specific to the protein. Pentelute and co-workers report an optimized protocol using their automated fast-flow peptide synthesis (AFPS) system that is capable of synthesizing full-length proteins of up to 164 amino acids, without the use of chemical ligation techniques, on time-scales that rival recombinant expression.


#

Comment

To synthesize full-length proteins, each step in the peptide coupling reaction had to be carefully optimized. The critical variables the authors identified included preactivation temperature and time, reaction temperature and Fmoc-deprotection conditions. The reaction coupling efficiency could be indirectly evaluated using an in-line UV/Vis spectrometer. AFPS was capable of synthesizing full-length proteins 10 times faster than traditional SPPS, with higher overall yield and purity. After purification and folding, the synthesized proteins possessed the same biological activity and biophysical characteristics as recombinant proteins. By using this technology, unnatural AAs can be easily incorporated and site-specific mutations are possible.


#
#

Publication History

Publication Date:
18 August 2020 (online)

© 2020. Thieme. All rights reserved

Georg Thieme Verlag KG
Rüdigerstraße 14, 70469 Stuttgart, Germany