Study On The Production Of Poly-?-glu-Tamic Acid For Bacillus Natto Induced By High Voltage Corona Electric Field

In order to improve the y-polyglutamic acid production ability,the Bacillus natto N-0 was mutagenized by a needle plate corona field,which can produce a high-yield strain N-E5.2 whose production of ?-PGA could reach 21.2 g·L-1 at the field condition of 3kV-30min-0.8cm conditions.That is to say the production of ?-PGA is eight times more than the initial strain N-0,after adding different kinds of rare earth elements in solid medium to mutageniz the strain N-E5.2b,?-PGA could reach 22.8 g·L-1,the production of ?-PGA increased 7.5%.The mutant strains was genetically stable in 7 generations.By using response surface methodology to optimize fermentation medium components that was the mutagenic B.natto N-E5.2b produced poly-?-glutamic acid.Building response equation to predict the optimal medium(g·L-1):peptone 15,sucrose 46.38,sodium glutamate 45.85,yeast extract powder 2.5,NaCl 5,MgSO4·7H2O 0.5,MnSO4·4H2O 0.4,CaCI2 0.25,K2HPO4·3H2O 5.2 and KH2PO4 2.At the same time,the production of ?-PGA could reach 38.94 g-L-1,which was 70.8%higher than before the optimization.The pgsBCA synthetase gene of six different treated strains was extracted and amplified,and the amino acid sequences of pgsB,pgsC and pgsA were compared with those of the synthetase proteins PgsB,PgsC and PgsA.The results showed that the pgsB,pgsC and pgsA genes contained about 1170,450 and 1140 nucleotides,respectively encoding about 390,150 and 380 amino acids.The high-voltage corona electric field and rare-earth element mutagenesis resulted in substitution and transversion,insertion and deletion in their nucleotide sequence of the ?-PGA synthase gene cluster pgsBCA,and the substitution,insertion and deletion in amino acids in their corresponding tamino acid sequence of the synthetase protein PgsBCA.There were a large effect on the start and end of the sequence.The PgsA enzyme protein changed significantly in the range of 30?40 amino acids.