Screening Of The Strain Producing Glutamic Acid Decarboxylase And Synthesis Of γ-aminobutyric Acid

γ-Aminobutyric acid is a non-protein amino acid that widelyexists inplants, animals andmicroorganisms. Researchers have found that γ-Aminobutyric acid has significantphysiological functions in cardiovascular disease, neurological disease, liver and kidney andregulation of the endocrine system. Therefore, it has a huge prospect in applications for drugtherapy and functional foods.Firstly, this study found6strains through screening process which could produce γ-aminobutyric acid by cell transformation from commercial yogurt and pickles. The glutamicacid decarboxylase activities of these6strains were measured to further screen out one strainthat had the highest decarboxylase activity. According to the strain morpholo gical,physiological, biochemical characteristics and16S rRNA gene sequence, the target strain wasfinally identified as Lactobacillus plantarum and was named as Lactobacillus plantarumSK-30. The final product of the reaction, which was catalyzed by the enzyme fromLactobacillus plantarum SK-30, was analyzed and identified as γ-aminobutyric acid byLC/MSMS.Medium composition and culture conditions have a significant impact on the amount ofenzyme production of bacterias. Medium compositions and culture conditions were studiedaccording to decarboxylase activity on carbon source, nitrogen source, metal ions, sodiumglutamate, incubation temperature, pH, etc. The best medium composition and cultureconditions were determined by orthogonal experiment: the best carbon source was sucrose,concentration15g/L; the best nitrogen source was the complex nitrogen, concentration25g/L;added metal ions were Zn2+and Mn2+, concentration was1mmol/L; sodium glutamateconcentration10g/L, culture temperature30C, initial pH6.2, incubation time20h. Glutamicacid decarboxylase activity was33.02U/L under the conditions before.This paper also studied the effects of the permeability conditions on glutamatedecarboxylase activity for the first time, specifically the permeability reagent, permeabilityreagent concentration, permeability treatment temperature and permeability treatment timeThe optimal processing conditions are determined as belows: permeability reagent was0.4%Triton X-100, the treatment time was10min, the treatment temperature was20C. This paperalso studied how the temperature and pH affect glutamate decarboxylase activity, and foundthat optimum temperature dropped to30C and the optimum pH was still4.2afterpermeability treatment. The thermal stability of the glutamic acid decarboxylase afterpermeability treatment was also studied; the result indicated a good stability after permeability treatment. Enzyme reaction kinetic studies showed that the enzyme reaction wasmainly affected by the internal diffusion, and external diffusion was largely independentThis paper also explored the effects of the reaction conditions to γ-aminobutyric acidcontent produced by Lactobacillus plantarum SK-30, specifically the cell concentration,substrate concentration, buffer solution concentration and reaction time. The optimumconditions are determined: cell concentration was120g/L, the substrate concentration was10g/L, buffer solution concentration was0.2mol/L, and reaction time was12h. The γ-aminoacid yield was4.0g/L under the condition before, which is higher than our laboratory strainsused in industrial production under the same condition (3.8g/L). This paper also comparedthe permeability cell and intact cells on bioconversion reaction time curve, and found that thepermeability can significantly shorten the reaction time and slightly improve γ-amino acidyield. This means that permeability technology also has the industrial application prospect...