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Local encoding of computationally designed enzyme activity.

Publication ,  Journal Article
Allert, M; Dwyer, MA; Hellinga, HW
Published in: J Mol Biol
February 23, 2007

One aim of computational protein design is to introduce novel enzyme activity into proteins of known structure by predicting mutations that stabilize transition states. Previously, we showed that it is possible to introduce triose phosphate isomerase activity into the ribose-binding protein of Escherichia coli by constructing 17 mutations in the first two layers of residues that surround the wild-type ligand-binding site. Here, we report that these mutations can be "transplanted" into a homologous ribose-binding protein, isolated from the hyperthermophilic bacterium Thermoanaerobacter tengcongensis, with retention of catalytic activity, substrate affinity, and reaction pH dependence. The observed 10(5)-10(6)-fold rate enhancement corresponds to 70% of the maximally known transition-state binding energy. The wild-type sequences in these two homologues are almost perfectly conserved in the vicinity of their ribose-binding sites, but diverge significantly at increasing distance from these sites. The results demonstrate that the computationally designed mutations are sufficient to encode the observed enzyme activity, that all the observed activity is encoded locally within the layer of residues directly in contact with the substrate and that, in this case, at least 70% of transition state stabilization energy can be achieved using straightforward considerations of stereochemical complementarity between enzyme and reactants.

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Published In

J Mol Biol

DOI

ISSN

0022-2836

Publication Date

February 23, 2007

Volume

366

Issue

3

Start / End Page

945 / 953

Location

Netherlands

Related Subject Headings

  • Triose-Phosphate Isomerase
  • Thermoanaerobacter
  • Software
  • Sequence Homology, Amino Acid
  • Protein Structure, Secondary
  • Protein Engineering
  • Periplasmic Binding Proteins
  • Molecular Sequence Data
  • Kinetics
  • Hydrogen-Ion Concentration
 

Citation

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Allert, M., Dwyer, M. A., & Hellinga, H. W. (2007). Local encoding of computationally designed enzyme activity. J Mol Biol, 366(3), 945–953. https://doi.org/10.1016/j.jmb.2006.12.002
Allert, Malin, Mary A. Dwyer, and Homme W. Hellinga. “Local encoding of computationally designed enzyme activity.J Mol Biol 366, no. 3 (February 23, 2007): 945–53. https://doi.org/10.1016/j.jmb.2006.12.002.
Allert M, Dwyer MA, Hellinga HW. Local encoding of computationally designed enzyme activity. J Mol Biol. 2007 Feb 23;366(3):945–53.
Allert, Malin, et al. “Local encoding of computationally designed enzyme activity.J Mol Biol, vol. 366, no. 3, Feb. 2007, pp. 945–53. Pubmed, doi:10.1016/j.jmb.2006.12.002.
Allert M, Dwyer MA, Hellinga HW. Local encoding of computationally designed enzyme activity. J Mol Biol. 2007 Feb 23;366(3):945–953.
Journal cover image

Published In

J Mol Biol

DOI

ISSN

0022-2836

Publication Date

February 23, 2007

Volume

366

Issue

3

Start / End Page

945 / 953

Location

Netherlands

Related Subject Headings

  • Triose-Phosphate Isomerase
  • Thermoanaerobacter
  • Software
  • Sequence Homology, Amino Acid
  • Protein Structure, Secondary
  • Protein Engineering
  • Periplasmic Binding Proteins
  • Molecular Sequence Data
  • Kinetics
  • Hydrogen-Ion Concentration