Optimization Of GABA Production Of Streptococcus Thermophilus And The Effect Of The PrtS Gene On GABA Production

GABA(?-aminobutyric acid)is an important bioactive substance.GABA has antianxiety,anti-depression,and other physiological functions,widely exists in animals,plants and microorganisms.Recently,screening of GABA-producing lactic acid bacteria(LAB)has become popular.In order to obtain excellent strains with high GABA production,we studied the GABA production of S.thermophilus CS9,CS11,CS12,CS17,CS19,and CS22.The PCR production showed six strains of S.thermophilus contain gad gene demonstrated that six strains are GABA producing bacteria.This study focused on the ability of the strain to produce GABA in fermented milk.We used three methods to extract protein from fermented milk;among them,the normal method is the best.The pH value is a key factor affecting the synthesis of GABA.In this study,the effects of different pH values on the synthesis of GABA was analyzed.When the initial pH value was 4.5,GABA production decreased of all isolates.GABA yields of CS9,CS12 and CS19 strains were highest at pH 6.0,5.0,and 5.5,respectively.The above results indicated that low pH was not conducive to the synthesis of GABA by lactic acid bacteria,and the pH value corresponding to the optimal yield of GABA by different strains was different.Adding an appropriate of monosodium glutamate(MSG)is beneficial to the synthesis of GABA.When 4% of MSG,CS19 strain produce GABA yield reach up to 11 453.17±164.80 mg/L.CS9 produces the highest GABA yield reached 2 181.24±112.24 mg/L with 2% MSG supplemented.The highest GABA yield of CS12 strain was 1 987.98±35.52 mg/L in medium supplemented with 1.5% MSG.Pyridoxal-5 '-phosphate(PLP)is a cofactor in the synthesis of GAD,a key enzyme in GABA,which can effectively promote MSG convert into GABA.CS19 was fermented at 72 hours in the medium with 150 ?m of PLP,the highest GABA yield reached 10 898.43±310.79 mg/L.CS9 was fermented in medium with 50 ?m of PLP at 120 hours,and the GABA content reached 1 183.08 ±76.43 mg/L.S.thermophilus CS11,CS12,CS17,and CS19 contained prtS gene while CS9 and CS22 do not have prtS genes.Through phylogenetic tree construction showed PrtS have an ancestor closely to Prt B than Prt P.From the physiological cultured in skim milk confirmed,CS12 and CS19 had rapid growth capacity than CS9 results in CS19,and CS12 have a proteolytic system than CS9.This study showed that PrtS was beneficial to hydrolysis protein to supported acidification in milk but had no impact on GABA production.In the future,the more prolongation of fermentation time with higher MSG percentage should be studied,and the study role of prtS gene on GABA production in different strains should be done.