Influence of reaction conditions on the enantioselectivity of biocatalyzed C-C bond formations under high pressure conditions.

Benzoylformate decarboxylase (BFD, EC 4.1.1.

Influence of the hydrostatic pressure and pH on the asymmetric 2-hydroxyketone formation catalyzed by Pseudomonas putida benzoylformate decarboxylase and variants thereof.

Benzoylformate decarboxylase (BFD) from Pseudomonas putida is a thiamine diphosphate-dependent (ThDP) enzyme that catalyzes the asymmetric C--C bond formation to (S)-2-hydroxypropiophenone [(S)-HPP] starting from benzaldehyde and acetaldehyde. The enantioselectivity of BFD was shown to be a function of temperature and substrate concentration. It can additionally be changed by site-directed mutagenesis on hot spot positions in the active site.

Structural and kinetic studies on native intermediates and an intermediate analogue in benzoylformate decarboxylase reveal a least motion mechanism with an unprecedented short-lived predecarboxylation intermediate.

The thiamin diphosphate- (ThDP-) dependent enzyme benzoylformate decarboxylase (BFDC) catalyzes the nonoxidative decarboxylation of benzoylformic acid to benzaldehyde and carbon dioxide. To date, no structural information for a cofactor-bound reaction intermediate in BFDC is available. For kinetic analysis, a chromophoric substrate analogue was employed that produces various absorbing intermediates during turnover but is a poor substrate with a 10(4)-fold compromised kcat.

Practical application of different enzymes immobilized on sepabeads.

The immobilization of an endoglucanase, benzoylformate decarboxylase (BFD) from Pseudomonas putida, as well as of lipase B from Candida antarctica (CALB) onto the carrier supports Sepabeads EC-EP, Sepabeads EC-EA, and Sepabeads EC-BU was accomplished. It is shown that via these immobilized biocatalysts the synthesis of both fine and bulk chemicals is possible. This is illustrated by the syntheses of polyglycerol esters and (S)-hydroxy phenyl propanone.

Diffusional fluorescence quenching of aromatic hydrocarbons.

The quenching of the fluorescence of five aromatic hydrocarbons by three halogenated organics and by molecular oxygen has been measured. Both fluorescence intensity and fluorescence lifetime measurements have been employed to validate results and interpretation; linear Stern-Volmer analyses are shown to apply throughout. The fluorescence quenching rate constant of molecular oxygen for the five aromatic hydrocarbons is essentially equivalent to the diffusion rate constant independent of the fluorophore excitation energy.