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4EEZ

Crystal Structure of Lactococcus lactis Alcohol Dehydrogenase variant RE1

Summary for 4EEZ
Entry DOI10.2210/pdb4eez/pdb
Related4EEX
DescriptorAlcohol dehydrogenase 1, ZINC ION, TETRAETHYLENE GLYCOL, ... (4 entities in total)
Functional Keywordsalcohol dehydrogenase, site-saturation mutagenesis, directed evolution, isobutyraldehyde, biofuel, oxidoreductase
Biological sourceLactococcus lactis subsp. lactis
Total number of polymer chains2
Total formula weight74142.16
Authors
Liu, X.,Bastian, S.,Snow, C.D.,Brustad, E.M.,Saleski, T.,Xu, J.H.,Meinhold, P.,Arnold, F.H. (deposition date: 2012-03-28, release date: 2012-09-26, Last modification date: 2024-02-28)
Primary citationLiu, X.,Bastian, S.,Snow, C.D.,Brustad, E.M.,Saleski, T.E.,Xu, J.H.,Meinhold, P.,Arnold, F.H.
Structure-guided engineering of Lactococcus lactis alcohol dehydrogenase LlAdhA for improved conversion of isobutyraldehyde to isobutanol.
J.Biotechnol., 164:188-195, 2012
Cited by
PubMed Abstract: We have determined the X-ray crystal structures of the NADH-dependent alcohol dehydrogenase LlAdhA from Lactococcus lactis and its laboratory-evolved variant LlAdhA(RE1) at 1.9Å and 2.5Å resolution, respectively. LlAdhA(RE1), which contains three amino acid mutations (Y50F, I212T, and L264V), was engineered to increase the microbial production of isobutanol (2-methylpropan-1-ol) from isobutyraldehyde (2-methylpropanal). Structural comparison of LlAdhA and LlAdhA(RE1) indicates that the enhanced activity on isobutyraldehyde stems from increases in the protein's active site size, hydrophobicity, and substrate access. Further structure-guided mutagenesis generated a quadruple mutant (Y50F/N110S/I212T/L264V), whose KM for isobutyraldehyde is ∼17-fold lower and catalytic efficiency (kcat/KM) is ∼160-fold higher than wild-type LlAdhA. Combining detailed structural information and directed evolution, we have achieved significant improvements in non-native alcohol dehydrogenase activity that will facilitate the production of next-generation fuels such as isobutanol from renewable resources.
PubMed: 22974724
DOI: 10.1016/j.jbiotec.2012.08.008
PDB entries with the same primary citation
Experimental method
X-RAY DIFFRACTION (1.9 Å)
Structure validation

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