Chiral synthons by selective redox reactions catalysed by hitherto unknown enzymes present in resting microbial cells

Abstract

Resting cells of various strict anaerobic or anaerobically grown facultative microorganisms are useful biocatalysts for redox reactions in a preparat scale. This will be shown for clostridia and Proteus species. Electron donors may be hydrogen gas, the cathode of an electrochemical cell, often formate, but in some cases also carbon monoxide. In addition to well-known reactions catalysed by yeasts, hitherto unknown reactions are also catalysed very effectively. These reactions are performed with substrate concentrations between 0.1-0.6 M. The productivity numbers are usually 10–500 times higher than with yeasts. For maximum productivity and long stability of up to 600 h under operational conditions, artificial electron mediators such as viologens or quinones and others in 1 mM concentration are essential. The stereo- and regioselectivity is excellent. In a few cases special conditions have to be observed which can be understood biochemically. The enzymes catalysing the reductions of many different 2-enoates, 2-enals, the oxo group of 2-oxo carboxylates, carboxylates without activation to aldehydes have been isolated and partially characterized. The enzymes are reversible and are not dependent on pyridine nucleotides. They are able to transfer single electrons. For dehydrogenations very different electron mediators can be used. For example anthraquinone-2,6-disulphonate is very effective. But useful NAD(P)H dependent enzymes are also present in clostridia. Some of them contain high enzyme activities by which NAD + or NADP + can be reduced with the above mentioned electron donors and mediators to NAD(P)H. Because these enzymes are reversible, selective NAD(P) + dependent dehydrogenations are also possible. By reduction many carboxylates carrying a chiral carbon atom in α-and/or in -position, (R)- hydroxy carboxylates especially those with additional functional groups and chiral alcohols have been prepared. 2-Oxo aldonates and aldarates and other 2-oxo carboxylates have been obtained by dehydrogenation. Preparative redox reactions under pyridine nucleotide regenerating conditions will be demonstrated with different substrates. Since all reaction sequences contain single electron transferring steps they can be carried out in electrochemical cells. Their over-all reaction rate can be continuously monitored by measuring the electrical current. The systems are very useful for the preparation of stereospecifically deuterated products. The optimal growth conditions of the commercially available microbial cells are indicated.