Abstract
BackgroundMandelic acid (MA), an important component in pharmaceutical syntheses, is currently produced exclusively via petrochemical processes. Growing concerns over the environment and fossil energy costs have inspired a quest to develop alternative routes to MA using renewable resources. Herein we report the first direct route to optically pure MA from glucose via genetic modification of the L-phenylalanine pathway in E. coli.ResultsThe introduction of hydroxymandelate synthase (HmaS) from Amycolatopsis orientalis into E. coli led to a yield of 0.092 g/L S-MA. By combined deletion of competing pathways, further optimization of S-MA production was achieved, and the yield reached 0.74 g/L within 24 h. To produce R-MA, hydroxymandelate oxidase (Hmo) from Streptomyces coelicolor and D-mandelate dehydrogenase (DMD) from Rhodotorula graminis were co-expressed in an S-MA-producing strain, and the resulting strain was capable of producing 0.68 g/L R-MA. Finally, phenylpyruvate feeding experiments suggest that HmaS is a potential bottleneck to further improvement in yields.ConclusionsWe have constructed E. coli strains that successfully accomplished the production of S- and R-MA directly from glucose. Our work provides the first example of the completely fermentative production of S- and R-MA from renewable feedstock.
Highlights
Mandelic acid (MA), an important component in pharmaceutical syntheses, is currently produced exclusively via petrochemical processes
Production of MA in the reaction broth was identified by HPLC equipped with a chiral column, and the results illustrated that hydroxymandelate synthase (HmaS) was able to convert phenylpyruvate to S-MA (Additional file 1), but not for the control (Additional file 1)
Trace amounts of R-MA were detected as well, the results revealing that HmaS is not a rigorous stereoinverting enzyme for catalyzing phenylpyruvate to S-MA
Summary
Mandelic acid (MA), an important component in pharmaceutical syntheses, is currently produced exclusively via petrochemical processes. Growing concerns over the environment and fossil energy costs have inspired a quest to develop alternative routes to MA using renewable resources. It has been reported that its stereoisomers can be prepared via enzymatic routes; for example, stereospecific nitrilases are used to produce S- or R-MA [10,11,12,13] and enantioselective microbes are employed to degrade S-MA or R-MA to yield R- or S-enantiomers [14,15,16], respectively. Pseudomonas polycolor and Micrococcus freudenreichii has been explored for the production of R-MA from the racemate [17] These commodity chemicals are manufactured entirely from petroleum-based feedstock such as benzaldehyde and mandelonitrile. Growing concerns over the environment and fossil resources costs have inspired a quest to develop more sustainable processes that afford these products from renewable feedstock at lower cost. Biological processes based on renewable resources such as glucose that would provide direct one-step production of chiral MA in a microbial fermentation process would be of commercial interest
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