降解大豆胰蛋白酶抑制剂的短小芽孢杆菌菌株及其胞外蛋白的鉴定  

Identification of Soybean Trypsin Inhibitor-degrading Bacillus pumilus Strains and Their Extracellular Proteins

作　　者：刘家维 黄昆仑[1] 梁志宏[1] LIU Jia-wei;HUANG Kun-lun;LIANG Zhi-hong(Beijing Advanced Innovation Center for Food Nutrition and Human Health,College of Food Science and Nutritional Engineering,China Agricultural University,Beijing 100048,China)

机构地区：[1]中国农业大学北京食品营养与人类健康高精尖创新中心,食品科学与营养工程学院,北京100048

出　　处：《现代食品科技》2020年第2期129-136,84,共9页Modern Food Science and Technology

基　　金：国家重点研发计划项目(2017YFC1600901)。

摘　　要：本研究以大豆胰蛋白酶抑制剂为唯一碳源,从豆类内部筛选获得3株具有降解大豆胰蛋白酶抑制剂的细菌。通过对菌株的16Sr DNA基因的分析,3株细菌分别被鉴定为枯草芽孢杆菌LZ013-1、短小芽孢杆菌LZ013-2和类芽孢杆菌LZ013-3。进一步研究发现LZ013-2菌株的上清液具有高效降解大豆胰蛋白酶抑制剂的活性,2h可将胰蛋白酶抑制剂抑制率降低73.60%。将LZ013-1和LZ013-2菌株应用于豆粕发酵中,两株芽孢杆菌均能高效降解胰蛋白酶抑制剂。原始豆粕的胰蛋白酶抑制剂活性为22.26 mg/g,经过LZ013-1和LZ013-2菌株的液态发酵后分别降低为0.50 mg/g和0.63 mg/g,经过固态发酵后分别降低为1.06 mg/g和1.03 mg/g。通过硫酸铵分级沉淀和凝胶柱层析,分离LZ013-2菌株发酵上清液中具有降解活性的蛋白质,并采用质谱鉴定分离得到的蛋白质,筛选并鉴定出2种活性蛋白,分别为肽酶S8(Uniprot ID:A0A2T0DB16)和肽酶M84(Uniprot ID:A8FIH7)。Soybean trypsin inhibitor(STI) was used as the sole carbon source, three strains of bacteria capable of degrading soybean trypsin inhibitors were isolated from the interior of soybeans. The 16 S rDNA sequence analysis revealed that the strains were Bacillus subtilis LZ013-1, Bacillus pumilus LZ013-2 and Paenibacillus sp. 013-3. Further studies showed that the supernatant of LZ013-2 exhibited a high activity for degrading soybean trypsin inhibitors, and the rate of trypsin inhibitor inhibition was reduced by 73.60% in 2 h. LZ013-1 and LZ013-2 were used in the fermentation of soybean meal, and were s found to degrade trypsin inhibitors efficiently. The trypsin inhibitor activity of the untreated soybean meal was 22.26 mg/g, which was reduced to 0.50 mg/g and 0.63 mg/g, respectively, after liquid fermentation with LZ013-1 and LZ013-2 strains, and decreased to 1.06 mg/g and 1.03 mg/g, respectively, after solid-state fermentation with LZ013-1 and LZ013-2 strains. The proteins with degrading activity were separated from the supernatant of the sample obtained via fermentation with LZ013-2 strain by ammonium sulfate fractionation and gel column chromatography. The separated proteins were identified by mass spectrometry. Two kinds of active proteins were screened and identified as peptidase S8(Uniprot ID: A0 A2 T0 DB16) and peptidase M84(Uniprot ID: A8 FIH7).

关 键 词：大豆胰蛋白酶抑制剂 芽孢杆菌 筛选 纯化 

分 类 号：TS201.2[轻工技术与工程—食品科学]