Related Experiment Video
Updated: Aug 16, 2026

Light-driven Enzymatic Decarboxylation
Published on: May 22, 2016
Directed evolution of the fatty-acid hydroxylase P450 BM-3 into an indole-hydroxylating catalyst
Q S Li1, U Schwaneberg, P Fischer
1Institut für Technische Biochemie, Universität Stuttgart, Germany. itbrsc@po.uni-stuttgart.de
Abstract:
The self-sufficient cytochrome P450 BM-3 enzyme from Bacillus megaterium catalyzes subterminal hydroxylation of saturated long-chain fatty acids and structurally related compounds. Since the primary structure of P450 BM-3 is homologous to that of mammalian P450 type II, it represents an excellent model for this family of enzymes. During studies on the directed evolution of P450 BM-3 into a medium-chain fatty-acid hydroxylase, several mutants, in particular the triple mutant Phe87Val, Leu188Gln, Ala74Gly, were observed to hydroxylate indole, producing indigo and indirubin at a catalytic efficiency of 1365 M(-1)s(-1) (kcat=2.73 s(-1) and Km=2.0 mM). Both products were unequivocally characterized by NMR and MS analysis. Wild-type P450 BM-3 is incapable to hydroxylate indole. These results demonstrate that an enzyme can be engineered to catalyze the transformation of substrates with structures widely divergent from those of its native substrate.
More Related Videos
Related Concept Videos
Overview of Fatty Acid Metabolism
Fatty acids are catabolized in a process called beta-oxidation, which takes place in the matrix of the mitochondria and converts their fatty acid chains into two-carbon units of acetyl groups. The acetyl...
Reactions of Aldehydes and Ketones: Baeyer–Villiger Oxidation
The carbonyl center is activated by...
Carboxylic Acids to Esters: Acid-Catalyzed (Fischer) Esterification Overview
Esters to Carboxylic Acids: Acid-Catalyzed Hydrolysis
During hydrolysis, the ester is first activated towards nucleophilic attack through the protonation of the carboxyl oxygen atom by the acid catalyst. The protonation makes the ester carbonyl carbon more electrophilic. In the next step, water acts as a nucleophile and adds to the...
Alkylation of β-Ketoester Enolates: Acetoacetic Ester Synthesis
Carboxylic Acids to Esters: Acid-Catalyzed (Fischer) Esterification Mechanism

