Biosynthesis of functional polyhydroxyalkanoates by engineered Halomonas bluephagenesis
Polyhydroxyalkanoates (PHA) have found widespread medical applications due to their biocompatibility and biodegradability , while further chemical modification requires functional groups on PHA. Halomonas bluephagenesis , a non-model halophilic bacterium serving as a chassis for the Next Generation Industrial Biotechnology (NGIB), was successfully engineered to express heterologous PHA synthase (PhaC) and enoyl coenzyme-A hydratase (PhaJ) from Aeromonas hydrophila 4AK4, along with a deletion of its native phaC gene to synthesize the short chain- co -medium chain-length PHA copolymers , namely poly(3-hydroxybutyrate- co -3-hydroxyhexanoate), poly(3-hydroxybutyrate- co -3-hydroxyhex-5-enoate) and poly(3-hydroxybutyrate- co -3-hydroxyhexanoate- co -3-hydroxyhex-5-enoate). After optimizations of the expression cassette and ribosomal binding site combined with introduction of endogenous acyl-CoA synthetase ( fadD ), the resulting recombinant strain H. bluephagenesis TDR4 achieved a remarkably high 3-hydroxyhexenoate (3HHxE) molar ratio of 35% when grown on glucose and 5-hexenoic acid as co-substrates. The total ratio of side chain consisting of 3HHx and 3HHxE monomers in the terpolymer can approach 44 mol%. H. bluephagenesis TDR4 was grown to a cell dry mass (CDM) of 30 g/L containing approximately 20% poly(3-hydroxybutyrate- co -22.75 mol% 3-hydroxy-5-hexenoate) in a 48-h of open and unsterile fermentation with a 5-hexenoic acid conversion efficiency of 91%. The resulted functional PHA containing 12.5 mol% 3-hydroxy-5-hexenoate exhibits more than 1000% elongation at break. The engineered H. bluephagenesis TDR4 can be used as an experimental platform to produce functional PHA.
危险品化学品经营许可证（带存储）