Alexandra N. E. Weissfloch

Bio: Alexandra N. E. Weissfloch is an academic researcher from McGill University. The author has contributed to research in topics: Lipase & Candida rugosa. The author has an hindex of 4, co-authored 6 publications receiving 1070 citations.

A structure-based rationalization of the enantiopreference of subtilisin toward secondary alcohols and isosteric primary amines

Abstract: Lipases favor one enantiomer of secondary alcohols (HOCHRR′) and isosteric primary amines (NH 2 CHRR′) while subtilisin favors the other enantiomer. In both cases, simple rules based on the size of the substituents at the stereocenter predict which enantiomer reacts faster. Thus, lipases and subtilisin are a pair of complementary enantioselective reagents for organic synthesis. The success of these rules suggests that these hydrolases distinguish between enantiomer primarily by the size of the substituents. Previously, we proposed a molecular mechanism for the enantiopreference of lipases based on the X-ray crystal structure of transition state analogs bound to a lipase. Here we suggest that a similar mechanism can also account for the opposite enantiopreference of subtilisin. The catalytic machinery (catalytic triad plus the oxyanion-stabilizing residues) in lipases is approximately the mirror image of that in subtilisin. In both hydrolases, the protein fold, as it assembles the catalytic machinery, also creates a restricted pocket for one substituent in the substrate (‘M’ or medium-sized). However, the catalytic His residue lies on opposite sides of this pocket in the two hydrolases. We propose that enantioselection arises from (1) the limited size of this pocket, (2) and a required hydrogen bond between the catalytic His and the oxygen or nitrogen of the alcohol or amine. This mechanism for enantioselection differs from that proposed by Derewenda and Wei who focussed on which carbonyl face in the ester or amide is attacked. Lipases and subtilisin indeed attack opposite faces, but we propose that this difference does not set the enantiopreference toward secondary alcohols.

Bacterial Biocatalysts: Molecular Biology, Three-Dimensional Structures, and Biotechnological Applications of Lipases

Abstract: ▪ Abstract Bacteria produce and secrete lipases, which can catalyze both the hydrolysis and the synthesis of long-chain acylglycerols. These reactions usually proceed with high regioselectivity and enantioselectivity, and, therefore, lipases have become very important stereoselective biocatalysts used in organic chemistry. High-level production of these biocatalysts requires the understanding of the mechanisms underlying gene expression, folding, and secretion. Transcription of lipase genes may be regulated by quorum sensing and two-component systems; secretion can proceed either via the Sec-dependent general secretory pathway or via ABC transporters. In addition, some lipases need folding catalysts such as the lipase-specific foldases and disulfide-bond–forming proteins to achieve a secretion-competent conformation. Three-dimensional structures of bacterial lipases were solved to understand the catalytic mechanism of lipase reactions. Structural characteristics include an α/β hydrolase fold, a catalytic ...