摘要
从传统泡菜中分离得到16株乳杆菌株,通过脱脂乳平板试验,从中初步筛选出12株产蛋白酶的菌株,通过复筛得到1株高产蛋白酶菌株L4。采用PCR技术对该菌株进行鉴定,通过硫酸铵盐析、透析袋透析收集胞外产物,以交联葡聚糖凝胶G-75层析纯化蛋白酶,以SDS-PAGE电泳确定蛋白酶分子量,同时对不同条件下蛋白酶的酶学特性进行研究。结果表明,该菌株的碱基序列与多株植物乳杆菌相应的16S r DNA序列99%相似,鉴定其为植物乳杆菌;蛋白酶分子量为40 k Da,在p H为9时胞外蛋白酶活性最高,其最适温度为50~60℃。酶活抑制试验显示Ca2+、Ba2+、Zn2+、EDTA能大大抑制该蛋白酶活性,而Mg2+、Mn2+、Cu2+、PMSF对胞外蛋白酶活力影响不大。
16 Lactobacillus strains were isolated from traditional pickled vegetable and initially screened to obtain12 protease-producing strains by the skimmed milk treadmill test. A strain with high productivity of protease,namely L4,was obtained after rescreening. PCR was performed to identify this strain. The extracellular products were collected by salting out with ammonium sulfate,followed by dialysis. The protease was purified by Sephadex G-75. The results indicated that the 16 S r DNA nucleotide sequence of this strain had 99% sequence identity to the corresponding sequences of several Lactobacillus strains,suggesting that L4 could be identified as Lactobacillus plantarum. Molecular weight of the protease was about 40 k Da as determined by SDS-PAGE. Investigation on the enzymology characteristics indicated that the enzyme exhibited the highest activity at p H9. 0 and the optimum temperature was 50- 60 ℃. The extracellular protease was greatly inhibited by Ca2 +,Ba2 +,Zn2 +,and EDTA,but was not inhibited by Mg2 +,Mn2 +,Cu2 +,and PMSF.
出处
《食品与发酵工业》
CAS
CSCD
北大核心
2015年第6期35-40,共6页
Food and Fermentation Industries
基金
浙江医学高等专科学校科研基金(项目编号2015B05)
