Breeding of useful microbes for degradation of cholesterol in food materials

为了降解食品原料中的胆固醇，我们尝试开发出具有降解胆固醇新功能的乳酸菌。三年来，我们致力于以下研究工作，取得了丰硕的成果。从日本、印度尼西亚、菲律宾、泰国、尼泊尔、越南、蒙古和埃及的传统发酵食品中分离到许多乳酸菌菌株。从菲律宾发酵水产品“布荣班古斯”中分离到一株植物乳杆菌，并对其进行了质粒分析。利用植物乳杆菌的质粒片段，构建了在乳酸菌和大肠杆菌中复制的两个有用的载体。利用所构建的载体，获得了转化植物乳杆菌的最佳条件，并建立了乳酸菌的宿主载体系统。将胆固醇氧化酶基因(ChoA)与载体重组，转化植物乳杆菌菌株Choa^+.5。对编码胆固醇降解酶的基因进行克隆和分析。发现了编码酮类固醇-Delta、I-脱氢酶、酮类-Delta、5-异构酶及其调控因子的基因。这些胆固醇降解酶正计划在乳酸菌中表达。这些含有胆固醇的食品原料将被这些工程乳酸菌处理。

For the purpose of degradation of cholesterol in foods materials, we have tried to bread lactic acid bacteria having a new function to degrade cholesterol. For three years, we have devoted the following research work and obtained fruitful results.1. Many strains of lactic acid bacteria from traditional fermented foods in Japan, Indonesia, Philippines, Thailand, Nepal, Vietnam, Mongolia, and Egypt were isolated.2. A strain of Lactobacillus plantarum isolated from a fermented fishery product, "Burong Bangus" in Philippines was selected and analyzed plasmids. By using the fragment of plasmids from L.plantarum, two useful vectors replicated in lactic acid bacteria and Escherichia coli were constructed.3. By using the vector constructed, we obtained the best conditions of transformation of L.plantarum strain and established host-vector system in lactic acid bacteria.4. The cholesterol oxidase gene (choA) was recombined with the vector and the strain of L.plantarum was transformed to choA^+.5. The genes encoded cholesterol degradating enzymes were cloned and analyzed. The genes encoded ketosteroid-DELTA^I-dehydrogenase, ketosteroid-DELTA^5-isomerase and the regulator were found.6. These cholesterol degradation enzymes are planning to express in lactic acid bacteria. The cholesterol containing food materials will be treated by these engineered lactic acid bacteria.