Breeding of useful microbes for degradation of cholesterol in food materials

为了降解食品材料中的胆固醇，我们尝试了具有降解胆固醇的新功能的乳酸菌。三年来，我们致力于以下研究工作，并取得了丰硕的成果。从日本、印度尼西亚、菲律宾、泰国、尼泊尔、越南、蒙古、埃及等国家的传统发酵食品中分离到多株乳酸菌.从菲律宾的一种发酵水产品“Burong Bangus”中分离到一株植物乳杆菌，对其进行质粒分析。利用植物乳杆菌的质粒片段，构建了两个在乳酸菌和大肠杆菌中复制的载体.利用构建的载体，获得了植物乳杆菌转化的最佳条件，并建立了乳酸菌宿主-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^+.对编码胆固醇降解酶的基因进行了克隆和分析。发现了编码甾酮-DELTA ^I-脱氢酶、甾酮-DELTA ^5-异构酶及其调节子的基因。这些胆固醇降解酶正计划在乳酸菌中表达。这些工程乳酸菌将处理含胆固醇的食品原料。

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.