5HQ3

Stable, high-expression variant of human acetylcholinesterase


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.60 Å
  • R-Value Free: 0.253 
  • R-Value Work: 0.201 
  • R-Value Observed: 0.204 

Starting Model: experimental
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wwPDB Validation   3D Report Full Report


This is version 1.5 of the entry. See complete history


Literature

Automated Structure- and Sequence-Based Design of Proteins for High Bacterial Expression and Stability.

Goldenzweig, A.Goldsmith, M.Hill, S.E.Gertman, O.Laurino, P.Ashani, Y.Dym, O.Unger, T.Albeck, S.Prilusky, J.Lieberman, R.L.Aharoni, A.Silman, I.Sussman, J.L.Tawfik, D.S.Fleishman, S.J.

(2016) Mol Cell 63: 337-346

  • DOI: https://doi.org/10.1016/j.molcel.2016.06.012
  • Primary Citation of Related Structures:  
    5HQ3

  • PubMed 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 ∼2,000-fold higher levels in E. coli compared to wild-type hAChE and exhibited 20°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.


  • Organizational Affiliation

    Department of Biomolecular Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Acetylcholinesterase548Homo sapiensMutation(s): 51 
Gene Names: ACHE
EC: 3.1.1.7
UniProt & NIH Common Fund Data Resources
Find proteins for P22303 (Homo sapiens)
Explore P22303 
Go to UniProtKB:  P22303
PHAROS:  P22303
GTEx:  ENSG00000087085 
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP22303
Sequence Annotations
Expand
  • Reference Sequence
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 2
MoleculeChains Sequence LengthOrganismDetailsImage
Acetylcholinesterase549Homo sapiensMutation(s): 51 
Gene Names: ACHE
EC: 3.1.1.7
UniProt & NIH Common Fund Data Resources
Find proteins for P22303 (Homo sapiens)
Explore P22303 
Go to UniProtKB:  P22303
PHAROS:  P22303
GTEx:  ENSG00000087085 
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP22303
Sequence Annotations
Expand
  • Reference Sequence
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.60 Å
  • R-Value Free: 0.253 
  • R-Value Work: 0.201 
  • R-Value Observed: 0.204 
  • Space Group: P 43 21 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 89.534α = 90
b = 89.534β = 90
c = 395.305γ = 90
Software Package:
Software NamePurpose
REFMACrefinement
MOSFLMdata reduction
SCALAdata scaling
PHASERphasing

Structure Validation

View Full Validation Report



Entry History 

Revision History  (Full details and data files)

  • Version 1.0: 2016-07-27
    Type: Initial release
  • Version 1.1: 2016-08-03
    Changes: Database references
  • Version 1.2: 2017-08-23
    Changes: Structure summary
  • Version 1.3: 2017-11-08
    Changes: Database references, Source and taxonomy, Structure summary
  • Version 1.4: 2024-01-10
    Changes: Data collection, Database references, Refinement description
  • Version 1.5: 2024-10-23
    Changes: Structure summary