title: Alternative fermentation pathway of cinnamic acid production via phenyllactic acid.
abstract: Cinnamic acid (CA) is the chemical basis for bulk production of flavoring reagents and chemical intermediates, and it can be fermented from biomass. Phenylalanine ammonia lyase (PAL) has been used exclusively in the bacterial fermentation of sugar biomass in which the fermentation intermediate phenylalanine is deaminated to CA. Here, we designed an alternative metabolic pathway for fermenting glucose to CA. An Escherichia coli strain that generates phenylalanine in this pathway also produces Wickerhamia fluorescens phenylpyruvate reductase and ferments glucose to D-phenyllactate (D-PhLA) (Fujita et al. Appl Microbiol Biotechnol 97: 8887-8894, 2013). Thereafter, phenyllactate dehydratase encoded by fldABCI genes in Clostridium sporogenes converts the resulting D-PhLA into CA. The phenyllactate dehydratase expressed by fldABCI in the D-PhLA-producing bacterium fermented glucose to CA, but D-PhLA fermentation and phenyllactate dehydration were aerobic and anaerobic processes, respectively, which disrupted high-yield CA fermentation in single batch cultures. We overcame this disruption by sequentially culturing the two strains under aerobic and anaerobic conditions. We optimized the incubation periods of the respective aeration steps to produce 1.7 g/L CA from glucose, which exceeded the yield from PAL-dependent glucose fermentation to CA 11-fold. This process is a novel, efficient alternative to conventional PAL-dependent CA production.
authors.affiliation: Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, 305-8572, Japan.
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Masuo
authors.affiliation: Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, 305-8572, Japan.
authors.forename: Yuta
authors.initials: Y
authors.lastname: Kobayashi
authors.affiliation: Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, 305-8572, Japan.
authors.forename: Ken-Ichi
authors.initials: K
authors.lastname: Oinuma
authors.affiliation: Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, 305-8572, Japan. takaya.naoki.ge@u.tsukuba.ac.jp.
authors.forename: Naoki
authors.initials: N
authors.lastname: Takaya
mesh_terms: D000332:Aerobiosis; D000693:Anaerobiosis; D002934:Cinnamates / Q000378:metabolism*; D003013:Clostridium / Q000201:enzymology / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism*; D007773:Lactates / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D004718:Saccharomycetales / Q000201:enzymology / Q000235:genetics
chemical_list: D002934:Cinnamates; D007773:Lactates; D011994:Recombinant Proteins; C029010:cinnamic acid; C017648:3-phenyllactic acid; D005947:Glucose
keywords: Cinnamic acid; Clostridium sporogenes; Escherichia coli; Phenyllactate dehydratase; Phenylpyruvate reductase