Search

Synthetic enzyme cascades in living cells often lack efficiency owing to the formation of byproducts by endogenous enzymes or toxicity of the cascade intermediates. Highly reactive aldehyde species can trigger a metabolic stress response, and this leads to undesired side reactions and decreased yields. Owing to the metabolic background of Escherichia coli ( E. coli), aldehydes may be irreversibly oxidized to carboxylic acids or reduced to the corresponding alcohols. Herein, we applied an approach to equilibrate the aldehyde concentration in vivo. We oxidized primary alcohols to the corresponding aldehydes by AlkJ, an alcohol dehydrogenase from Pseudomonas putida. Introduction of a carboxylic acid reductase from Nocardia iowensis allowed the target compound to be retrieved from the carboxylate sink. Further reduction of the aldehydes to alcohols by endogenous E. coli enzymes completed the equilibration between alcohols, aldehydes, and carboxylic acids. Thus, the aldehyde concentrations remained below nonviable concentrations. We demonstrated the concept on several primary alcohols, which reached the redox equilibrium within 6 h and persisted up to 24 h. Subsequent combination with a dihydroxyacetone-dependent aldolase (Fsa1-A129S, E. coli) demonstrated that the reactive aldehyde species were freely available and gave the aldol product, (3 S,4 R)-1,3,4-trihydroxy-5-phenylpentan-2-one, in 70 % yield within short reaction times. [ABSTRACT FROM AUTHOR]

The Cover shows a biochemical cell factory (Escherichia coli) for the synthesis of chiral polyhydroxylated compounds via an artificial enzyme cascade.In their Communication, T. Bayer et al. presented a rational approach for tuning the concentration of toxic aldehyde intermediates in living cells. By combining an alcohol dehydrogenase and a carboxylic acid reductase, the redox equilibrium between alcohol, aldehyde and carboxylic acid species can be adjusted, thus maintaining the aldehyde concentration in a ‘hidden reservoir’ and therefore below the toxicity level, yet freely available for subsequent aldolase mediated reaction. More information can be found in the Communication by T. Bayer et al. on page 2919 in Issue 15, 2017 (DOI: 10.1002/cctc.201700469). [ABSTRACT FROM AUTHOR]