초록
<P><B>Abstract</B></P> <P>Adipic acid (AA) is an important dicarboxylic acid used for the manufacture of nylon and polyurethane plastics. In this study, a novel adipic acid biosynthetic pathway was designed by extending the <I>cis,cis</I>-muconic acid (MA) biosynthesis through biohydrogenation. Enoate reductase from <I>Clostridium acetobutylicum</I> (CaER), an oxygen-sensitive reductase, was demonstrated to have <I>in vivo</I> enzyme activity of converting <I>cis,cis</I>-muconic acid to adipic acid under microaerobic condition. Engineered <I>Escherichia coli</I> strains were constructed to express the whole pathway and accumulated 5.8 ± 0.9 mg/L adipic acid from simple carbon sources. Considering the different oxygen demands between <I>cis,cis</I>-muconic acid biosynthesis and its degradation, a co-culture system was constructed. To improve production, T7 promoter instead of <I>lac</I> promoter was used for higher level expression of the key enzyme CaER and the titer of adipic acid increased to 18.3 ± 0.6 mg/L. To decrease the oxygen supply to downstream strains expressing CaER, <I>Vitreoscilla</I> hemoglobin (VHb) was introduced to upstream strains for its ability on oxygen obtaining. This attempt further improved the production of this novel pathway and 27.6 ± 1.3 mg/L adipic acid was accumulated under microaerobic condition.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Construct a novel biosynthetic pathway of adipic acid <I>via</I> reduction of <I>cis,cis</I>-muconic acid. </LI> <LI> Employ an oxygen-sensitive enoate reductase for reducing <I>cis,cis</I>-muconic acid under microaerobic condition. </LI> <LI> Engineer a co-culture system to improve adipic acid production. </LI> <LI> Employ <I>Vitreoscilla</I> hemoglobin for balancing different oxygen demands of strains in co-culture system. </LI> </UL> </P>