保加利亚乳杆菌水解酪蛋白能力及产物分析

Study on the Ability of Lactobacillus delbrueckii subsp. bulgaricus to Hydrolyze Casein and Analysis of the Hydrolysate

摘要:

为了深入研究保加利亚乳杆菌酪蛋白水解情况。本试验主要以保加利亚乳杆菌ATCC 11842作为研究对象，同时以保加利亚乳杆菌KLDS 08007、KLDS 08009、KLDS 08010、KLDS 08014和KLDS 08018作为对照研究，测定不同菌株发酵液的酪蛋白水解度和多肽含量，同时对ATCC 11842的酪蛋白水解产物进行SDS-PAGE电泳分析和LC-MS分析。结果表明：ATCC 11842的水解能力最强，水解度为13.89%，多肽含量为0.63 mg/mL。测定六株保加利亚乳杆菌细胞壁蛋白酶的活性与水解能力进行比较分析发现，该酶活性与菌株的水解能力呈正相关，ATCC 11842的酶活力最强，达到15.52 U/mL，比活力为6.73 U/mg。通过电泳图谱发现，随着发酵时间的延长，酪蛋白逐渐水解，并生成3.4~21.0 ku的多肽。对水解物进行LC-MS分析，酪蛋白水解后共产生77种6~25肽的片段，其中6~7肽占片段种类的9%，8~14肽占多肽种类的77%。

Abstract:

In order to carry out an in-depth study on casein hydrolysis by Lactobacillus bulgaricus, Lactobacillus delbrueckii subsp. bulgaricus strain ATCC 11842 was used, with Lactobacillus delbrueckii subsp. bulgaricus strains KLDS 08007, KLDS 08009, KLDS 08010，KLDS 08014, and KLDS 08018 as controls. The degree of casein hydrolysis and peptide content of the fermentation broths from different strains were measured, and the casein hydrolysate of ATCC 11842 was further analyzed by sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and liquid chromatography-mass spectrometry (LC-MS). The results showed that ATCC 11842 had the strongest hydrolytic ability with the degree of hydrolysis as 13.89%, and the peptide content 0.63 mg/mL. The activities of cell envelope proteinases (CEPs) of the six strains were measured and compared with their hydrolytic abilities, and the results showed that their proteinase activity was positively related to their hydrolytic ability. ATCC 11842 exhibited the highest enzyme activity (up to 15.52 U/mL), and its specific enzyme activity was 6.73 U/mg. It can be seen from the electrophoretogram that the casein was hydrolyzed gradually with a prolonged fermentation time and 3.4~21.0 ku polypeptides were produced. As shown from the LC-MS analysis of the casein hydrolysate, 77 types of peptide fragments containing 6~25 residues were produced after casein hydrolysis, and the peptides containing 6~7 residues and 8~14 residues accounted for 9% and 77% of the total types of polypeptides, respectively.