Expression of genes regulated by acid adaptation related TCS1 of Lactobacillus delbrueckii subsp. bulgaricus improved acid tolerance of Lactococcus lactis subsp. cremoris NZ9000

Abstract:

[Objective] We investigated the acid tolerance mechanisms of Lactobacillus delbrueckii subsp. bulgaricus by heterologous expression of genes related to acid adaptation regulated by two-component system TCS1 (JN675228/JN675229) of Lb. bulgaricus in Lactococcus lactis subsp. cremoris NZ9000. [Methods] We used reverse transcription polymerase chain reaction and sodium dodecyl sulfate-polyacrylamide gel electrophoresis to verify the expression of acid adaptation related genes, such as adenine phosphoribosyltransferase (aprt), D-alanine-D-alanine ligase (ddl), oligopeptide ABC transporter (oppDII) and elongation factor Ts (tsf), regulated by TCS1 of Lb. bulgaricus, in Lc. lactis NZ9000. We validated acid treatment experiment to verify the effect of gene expression on acid stress tolerance of host bacteria. We confirmed the interactions between TCS1 and the expressed genes related to acid adaptation by yeast two hybrid. [Results] Our results show that aprt, ddl, oppDII and tsf were successfully expressed in Lc. lactis NZ9000. The expression of aprt and ddl genes increased the resistance of recombinant strains to acid stress by 75 and 114 times. The expression of oppDII and tsf genes had no significant effect on the acid resistance of the recombinant strains. Yeast two hybrid system showed that there was interaction between histidine protein kinase (HPK1) of TCS1 and Ddl, and HPK1-C domain was the key region of the interaction. [Conclusion] The acid tolerance of strains expressing Aprt and Ddl was significantly higher than that of the control strain. The results of this study can provide references for the strategies of obtaining acid resistance of Lb. bulgaricus and other similar strains.