%0 Journal Article
%A Oeggl, Reinhard
%A Neumann, Timo
%A Gätgens, Jochem
%A Romano, Diego
%A Noack, Stephan
%A Rother, Dörte
%T Citrate as Cost-Efficient NADPH Regenerating Agent
%J Frontiers in Bioengineering and Biotechnology
%V 6
%@ 2296-4185
%C Lausanne
%I Frontiers Media
%M FZJ-2018-07409
%P 196
%D 2018
%X The economically efficient utilization of NADPH or NADH-dependent enzymes requires the regeneration of consumed reduction equivalents. This cofactor regeneration is classically done via an additional substrate, and if necessary enzyme. We now demonstrate an easy-to-apply cofactor regeneration approach, which can especially be used in screening applications. Simply by applying citrate to a buffer or directly using citrate/-phosphate buffer NADPH can be regenerated by native enzymes of the TCA cycle, practically present in all aerobic living organisms. Apart from viable-culturable cells, this regeneration approach can also be applied with lyophilized cells and even crude cell extracts. This is exemplarily shown for the synthesis of 1 phenylethanol from acetophenone with several oxidoreductases. The mechanism of NADPH regeneration by TCA cycle enzymes was further investigated by a transient isotopic labeling experiment feeding [1,5-13C]citrate. This revealed that the regeneration mechanism can further be optimized by genetic modification of two competing internal citrate metabolization pathways, the glyoxylate shunt and the glutamate dehydrogenase.
%F PUB:(DE-HGF)16
%9 Journal Article
%$ pmid:30631764
%U <Go to ISI:>//WOS:000454341200001
%R 10.3389/fbioe.2018.00196
%U https://juser.fz-juelich.de/record/858538