王力源¹，秦 梦¹，刘春卯¹，赵 露¹，刘 博²，郑 翔^1*

（1.河北省微生物研究所有限公司，保定071051；2.保定学院，保定071000）

摘要：为探索一种价廉、无毒的乳糖代替异丙基-β-D巯基半乳糖苷（IPTG）作为诱导剂进行中性蛋白酶发酵的工艺，该研究以枯草芽孢杆菌工程菌（Bacillus Subtilis）WB800N/pHT43-npr 为试验菌株，对乳糖与IPTG两种诱导剂进行了发酵对比，在此基础上以乳糖作为诱导剂对菌株B. Subtilis WB800N/pHT43-npr 产中性蛋白酶发酵效果进行响应面优化。结果表明，添加15.00 g/L乳糖诱导剂与添加0.1% IPTG的诱导效果基本相同，对乳糖诱导方式进行响应面优化，获得最佳发酵条件为：培养温度35℃、发酵时间47h、初始pH值7.0、诱导剂添加时间为发酵3h时，最终所得蛋白酶活力为1 079.31 U/mL，与原有工艺IPTG诱导相比，酶活力提高了14.22%。在一定程度上降低了成本，去除了毒性，为中性蛋白酶的工业化应用提供了数据支撑。

关键词：枯草芽孢杆菌；乳糖诱导；蛋白酶；发酵工艺

中图分类号：Q939.97 文献标识码：A 文章编号：1674-506X（2024）05-0009-0008

Study on the Optimization of Protease Production Process of Bacillus subtilis Engineering Bacteria WB800N/pHT43-npr Induced by Lactose

WANG Liyuan¹，QIN Meng¹，LIU Chunmao¹，ZHAO Lu¹，LIU Bo²，ZHENG Xiang^1*

（1.Hebei Research Institute of Microbiology Co.，Ltd.，Baoding 071051，China；2.Baoding University，Baoding 071000，China）

Abstract：In order to explore a neutral protease fermentation process using low-cost, non-toxic lactose instead of IPTG as an inducer, this study used Bacillus subtilis engineering bacteria WB800N/pHT43-npr as the test strain, and compared the fermentation of lactose and IPTG inducers. On this basis, lactose was used as an inducer to optimize the fermentation effect of neutral protease produced by Bacillus subtilis engineering bacteria by response surface methodology. The results showed that the induction effect of adding 15.00 g/L lactose inducer was basically the same as that of adding 0.1% IPTG. The response surface methodology optimization of lactose induction method was carried out, obtaining the optimal fermentation conditions: culture temperature 35 ℃, fermentation time 47 h, initial pH 7.0, and adding inducers after 3 h fermentation. The final protease activity was 1 079.31 U/mL, which was 14.22 % higher than that induced by IPTG in the existing process. To a certain extent, it reduces the cost and removes the toxicity, which provides data support for the industrial application of neutral protease.

Keywords：Bacillus subtilis；lactose induction；protease；fermentation technology

doi：10.3969/j.issn.1674-506X.2024.05-002

收稿日期：2023-11-19

基金项目：河北省科学院基本科研业务费制度试点项目资金资助（2022PF02-2）；河北省科学院高层次人才培养与资助项目（2022Q2）

作者简介：王力源（1993-），男，硕士，助理研究员。研究方向：酶工程。

*通信作者：郑翔（1987-），男，博士，副研究员。研究方向：微生物学。

引用格式：王力源，秦梦，刘春卯，等.乳糖诱导枯草芽孢杆菌工程菌WB800N/pHT43-npr 产蛋白酶工艺优化研究［J］.食品与发酵科技，2024，60（5）：9-15，82.