Improved Viability and Proteome Analysis of Lactobacillus fermentum KLB12 upon Heat Stress

Lactobacillus fermentum KLB12의 열 전처리에 따른 열 스트레스 내성 증진 및 프로테옴 변화

In the previous study, we have isolated several vaginal lactobacilli from Korean woman and selected one of them (KLB12) for further study, which was indentified as Lactobacillus fermentum by sequence analysis of 16S rRNA gene. Formulated L. fermentum KLB12 can be used for ecological treatment of bacterial vaginosis. For pharmaceutical formulation, the spray-drying process is required where stress such as high temperature is routinely applied. In this study, we found that heat stress at 60$^{\circ}C$ for 20∼30min reduced the viable cell population of KLB12 by 10$\sub$6/～10$\sub$9/. However, adaptation of KLB12 cells at 52$^{\circ}C$ made them more thermotolerant upon exposure to 60$^{\circ}C$. The level of thermal protection in RSM (reconstituted skim milk) by adaptation in acid (pH 5), cold (4$^{\circ}C$), ethanol (3％), NaCI (0.3M) was also examined. Although not as efficient as the homologous stress, adaptations in both cold and NaCI gave considerable cross protection against heat stress. When chloramphenicol was added during heat adaptation, adaptation effect was abolished. This suggests that de novo protein synthesis is necessary during the adaptation process. Important changes in proteome during heat adaptation was examined with two-dimensional gel electrophoresis.

본 연구에서는 L. fermentum KLB12을 열 전처리 함으로서 제제화 과정 동안 거치게 되는 열 스트레스에 대한 내성이 증진됨을 확인하고, 최적의 열 전처리 조건을 수행하였다. 또한 열 전처리 뿐만 아니라, 저온과 열 전처리 조건에도 열 스트레스에 대한 간섭 효과를 확인하였다. 그리고 내성 증진에 신규 단백질 합성이 필요함을 확인하였으며 나아가, 2-D electrophoresis를 통하여 7개의 신규 단백질을 확인하였다. 따라서 이 균주를 제제화하기 위한 방법으로 열 전처리를 이용할 경우 생균력 유지에 큰 효과를 얻을 수 있다고 사료된다.