Related Experiment Video
Updated: May 18, 2026

A Toolkit to Enable Hydrocarbon Conversion in Aqueous Environments
Published on: October 2, 2012
Structure-guided engineering of Lactococcus lactis alcohol dehydrogenase LlAdhA for improved conversion of
Xiang Liu1, Sabine Bastian, Christopher D Snow
1Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
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.
Related Concept Videos
Crossed Aldol Reaction Using Strong Bases: Directed Aldol Reaction
Alcohols from Carbonyl Compounds: Reduction
Catalytic hydrogenation is similar to the reduction of an alkene or alkyne by adding H2 across the pi bond in the presence of transition metal catalysts like Raney Ni, Pd–C, Pt, or Ru. Aldehydes and ketones can be reduced by this method, often under mild to moderate heat (25–100°C) and...
Production of Organic Acids
Acid Halides to Alcohols: LiAlH4 Reduction
The mechanism proceeds in three steps. First, the nucleophilic hydride ion attacks the carbonyl carbon of the acid halide to form a tetrahedral intermediate. Next, the carbonyl group is re-formed, and the halide ion departs as a leaving group, generating an aldehyde. A second nucleophilic attack by the hydride yields an alkoxide ion, which, upon protonation, gives a primary alcohol as...
Base-Catalyzed Aldol Addition Reaction
Reactions of Aldehydes and Ketones: Baeyer–Villiger Oxidation
The carbonyl center is activated by...
