The Breeding Of Strains Producing Gamma-glutamyltranspeptidase

Theanine is the characteristics amino acid ,and also one of taste compounds. Theanine has a very important role in our daylife. However, conventional extraction methods, processes and procedures is very complicated and difficult to control,and extraction yield is very low.While the microbial transformation has been widely applied ,the method of synthesising theanine byγ-glutamyl peptide is more simple and effective. Currently the Microorganisms secretingγ-glutamyl peptidase mainly include Escherichia coli, Bacillus subtilis and Bacillus licheniformis strains and so on.But among the vast majority of strains,γ-glutamyl peptide is Intracellular enzymes. Datas show thatγ-glutamyl peptidase which Bacillus subtilis secretes can go through the cell membrane and cell wall to the broth,so it greatly simplifies the operation of the experiment.In this paper, highγ-glutamyl peptidase strain, and good genetic stability is Selected out, through their experiments by UV mutagenesis and Microwave irradiation on Bacillus subtilis which stored in the laboratory. The strains obtained by UV mutagenesis and Microwave irradiation have high stability and yield, and the activity ofγ-GGT are 3.56U/mL and 3.52U/mL.The experiment optimize the fermentation medium for Bacillus subtilis. According to the results, when the culture medium containing Glucose, yeast extract, pH7.5, and 60ml per 250ml , the activity ofγ-GGT is higer. By the orthogonal experimental design and optimization,we get the best combination of factors and levels,and the carbon source is glucose, nitrogen source for the corn flour, pH 8, with volume of 60ml.At the same time,γ-glutamyl glutamic enzymatic reaction has been studied in this experiment. The optimum temperature is 35℃, pH value of 7.5, and the metal ions Ca²⁺ and Mg²⁺have activation while Cu²⁺,Mn²⁺ and Zn²⁺ have inhibition on the activity ofγ-GGT.