Hill SE

References (1)

Title : Automated Structure- and Sequence-Based Design of Proteins for High Bacterial Expression and Stability - Goldenzweig_2016_Mol.Cell_63_337
Author(s) : Goldenzweig A , Goldsmith M , Hill SE , Gertman O , Laurino P , Ashani Y , Dym O , Unger T , Albeck S , Prilusky J , Lieberman RL , Aharoni A , Silman I , Sussman JL , Tawfik DS , Fleishman SJ
Ref : Mol Cell , 63 :337 , 2016
Abstract : Upon heterologous overexpression, many proteins misfold or aggregate, thus resulting in low functional yields. Human acetylcholinesterase (hAChE), an enzyme mediating synaptic transmission, is a typical case of a human protein that necessitates mammalian systems to obtain functional expression. We developed a computational strategy and designed an AChE variant bearing 51 mutations that improved core packing, surface polarity, and backbone rigidity. This variant expressed at approximately 2,000-fold higher levels in E. coli compared to wild-type hAChE and exhibited 20 degrees C higher thermostability with no change in enzymatic properties or in the active-site configuration as determined by crystallography. To demonstrate broad utility, we similarly designed four other human and bacterial proteins. Testing at most three designs per protein, we obtained enhanced stability and/or higher yields of soluble and active protein in E. coli. Our algorithm requires only a 3D structure and several dozen sequences of naturally occurring homologs, and is available at http://pross.weizmann.ac.il.
ESTHER : Goldenzweig_2016_Mol.Cell_63_337
PubMedSearch : Goldenzweig_2016_Mol.Cell_63_337
PubMedID: 27425410
Gene_locus related to this paper: human-ACHE