Enzymes in organic synthesis

Abstract

Several aspects of the use of enzymes as catalysts in organic synthesis have been explored. The stability of the enzyme glucose dehydrogenase useful in reduced nicotinamide cofactor (NAD(P)H) regeneration has been studied. The enzyme has been found to be much more stable than other enzymes commonly used in NAD(P)H regeneration. The enzyme FMN reductase has been used in oxidized nicotinamide cofactor (NAD(P)H) regeneration. The regeneration system using this enzyme is much faster and more efficient than the nonenzymatic system used previously. The substrate specificity of the enzyme hexokinase has been analyzed with regard to fluorinated analogs of D-glucose and analogs having an amino or thiol group at the five position. The enzyme has been found to accept a wide variety of analogs as substrates although the rates are often quite low. 3-Deoxy-3-fluoro-D-glucose has been converted to the 6-phosphate using hexokinase and ATP and this product has been converted chemically to 2-deoxy-2-fluoro-D-arabinose-5-phosphate. Fructose diphosphate aldolase in combination with glucose isomerase has been used in the synthesis of 6-deoxy-6-fluoro-D-glucose. Fluoro hydroxyacetone phosphate has been found to not be a substrate for aldolase but is a fairly good inhibitor. A seven carbon dideoxy ketose sugar has been synthesized using aldolase catalysis. Also 3,4-¹³C₂-D-glucose has been made from 1-¹³C-glyceraldehyde-3-phosphate using triosephosphate isomerase, fructose diphosphate aldolase and phosphoglucose isomerase as catalysts. Furyl carbinols have been prepared in high optical purity by reduction of the corresponding furyl ketones using the alcohol dehydrogenase from Thermoanaerobium brockii. Racemic furyl methyl carbinol has also been resolved to high optical purity by kinetic resolution using hydrolytic enzymes. Some compounds having low solubility in water have been prepared using enzymatic reductions in biphasic systems. The biphasic system allows the reaction to be carried out in a small reaction volume and reduces product inhibition. The use of arsenate and vanadate to stimulate activity of organic phosphate utilizing enzymes with their unphosphorylated substrate analogs has been studied. Several synthetically useful enzymes have been shown to demonstrate such activity. The utility of this approach to enzymatic synthesis has been demonstrated with the synthesis of a seven carbon sugar using dihydroxyacetone in the presence of arsenate as the substrate instead of dihydroxyacetone phosphate.