Reduced F₁F₀-ATPase Mutation Analysis of Lactobacillus Plantarum and Its Application in Sichuan Pickle as Probiotics Adjunct

Three mutation strains L. plantarum LPS08, LPS21 and LPS24 with reduced F₁F₀-ATPase activity were screened from acid production degenerationL. plantarum CCTCC 207202. In F₁F₀-ATPase β -subunit of LPS08, Phe-91, Ser-380 and Leu-436 were replaced by Leu, Pro and Pro, respectively. But these mutations do not markedly affect F₁F₀-ATPase activity, only lead to reduction of activity of 5.61%. In mutant strain LPS21, Leu-268 mutation has changed the F₁F₀-ATPase β-subunit, and thus it triggered a great change with 43.44% F₁F₀-ATPase activity reduction. In LPS24, the Asn-205, Ser-380 and Pro-419 were respectively replaced by Ser, Leu and Leu. For these mutations, F₁F₀-ATPase activity decreased 19.46%. In initial pH 6.5 MRS liquid medium, the pH values respectively mediated by mutation strain LPS08, LPS21 and LPS24 were 3.73, 3.68 and 4.25, and their acidities were 2.0 g/100 ml, 1.56 g/100 ml and 0.95g/100 ml after cultured at 37 ℃ for 72h. Compared with parent strain CCTCC 207202, only the mutation strain LPS21 could overcome post-acidification when their cultures were incorporated into aseptic Sichuan pickle products at 6 log CFU/ml salt brine level and kept at 30 ℃ up to 7 days, and its viable count in products(7.27 log CFU/ml) also satisfied the criteria for a probiotic food product. Therefore, the F₁F₀-ATPase mutant strain LPS21 is attractive as probiotics cultures adjunct in Sichuan pickle products to prevent post-acidification during the product handing, transportation and storage at ambient temperature.