The Study Of Producing Acrylic Acid By Biosynrhesis

Acrylic acid is a crucial chemical material, mainly produced from petroleum by chemical synthesis. However, this method is restricted as other chemical synthesis for their common shortcomings. Therefore, it is necessary to study and explore a biosynthesis pathway for producing acrylic acid.Acrylic acid is synthesized in two paths described as below in nature. The first way is to form the framework of propylene or acetate by changing the number of carbons before further modification. The other way is to form acrylic acid by reconstructing the existed tricarbon frame. In this work, we applied Alcaligenes faecalis, Rhodococcus erythropolis and Clostridium propionicum to realize the conversion of acrylic acid.By constructing the genome library of A. faecalis, DMSP Lyase gene was cloned and screened. The positive plasmids were to be expressed in E.coli BL21(DE3) for the conversion of acrylic acid from DMSP. However, mainly because of the capability of the genome library and screening, the positive strain was failed to obtain.Using the genome of R. erythropolis as the template, ami gene was successfully cloned and expressed in E.coli BL21(DE3). The expression mass, specific enzyme activity and the optimum reaction conditions of amidase were studied. The results showed there is a mount of inclusion body existed in E.coli BL21(DE3) after inducing and expressing ami gene. The optimum temperature of amidase was37℃with pH7.0. The specific enzyme activity of amidase could approximately reach2.8U/mg; the expression mass could nearly reach1.7mg/ml.In order to enhance the expression mass and enzyme activity, Bacillus subtilis was chose as the host. Five distinct promoters were selected for exploring the expression influence on ami gene. After comparing expression masses and enzyme activities, results displayed ami gene was expressed most effectively under the control of amyE promoter. The specific enzyme activity of amidase could reach8.7U/mg, with the expression mass of2.3mg/L. It is4times when compared with the wild strain Rhodococcus erythropolis. Under the optimum condition of37℃, pH7.0, the molar conversion of amidase could reach90%within6hr.The lcd gene and pct gene were both successfully cloned and expressed in C. propionicum o The recombinant C. propionicum was fermented in5L bioreactor. It was accomplished to synthesize acrylic acid from lactic acid. This work provided the basis for forthcoming research.