| 陈 丹， 游子娟， 黄伟谦，等.重组米曲霉丝氨酸羧肽酶O的性质鉴定及脱苦应用研究[J].广东农业科学,2015,42(5):103-108 |
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重组米曲霉丝氨酸羧肽酶O的性质鉴定及脱苦应用研究 |
| Characteristics of carboxypeptidase O from Asperigillus oryzae expressed in Pichia pastoris and its application in protein hydrolysis and debittering |
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| DOI： |
| 中文关键词: 米曲霉 重组羧肽酶 O 酶学性质 脱苦 |
| 英文关键词: Aspergillus oryzae recombinant carboxypeptidase O enzymatic characteristics debittering |
| 基金项目: |
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| 摘要点击次数: 1626 |
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| 中文摘要: |
| 为了研究羧肽酶在蛋白水解及对蛋白水解物脱苦中的重要作用‘利用毕赤酵母表达系统对米曲
霉（A sperigillus oryzae）羧肽酶 O（carboxypeptidaseO‘OcpO）进行表达。重组米曲霉羧肽酶 O（rOcpO）经过分子筛
和离子柱纯化‘对其进行脱糖基化及酶学性质的检测研究。 结果表明‘该蛋白酶是以糖蛋白的形式进行表达‘
分子量约为 74 ku‘脱糖基化后的相对分子质量约为 56 ku‘产量约为 20.4 nKat/mL。 该蛋白酶是一种酸性蛋白
酶‘最适 pH 值为 4.0‘且在 pH 3.0~6.0 时保持稳定；最适温度约为 40益‘具有良好的热稳定性‘在 60益孵育 1h
后仍能保持 63%的酶活；PMSF 能够显著抑制该酶酶活‘证明其为丝氨酸羧肽酶。 通过 rOcpO 对大豆蛋白和酪
蛋白的 Alcalase 蛋白酶水解物的进一步水解研究发现‘rOcpO 水解对两种水解液的苦味均有明显减轻‘并且水
解度有明显升高。 |
| 英文摘要: |
| In order to study the importance of carboxypeptidase in protein hydrolysis and debittering, a truncated
carboxypetidase O from Asperigillus oryzae was expressed in Pichia pastoris. The recombinant carboxypeptidase O
(rOcpO) was deglycosylated and characterized after purified through Sephadex G-75 and DEAE anionic-exchange
chromatography. The results showed that rOcpO was glycosylated of a molecular weight of about 74 ku and 56 ku
after been deglycosylated, the enzyme yield was about 20.4 nKat/mL. rOcpO was an acid protease, its optimal pH was
about 4.0, and it was stable in the pH range of 3-6; its optimal temperature was about 40益, it retained 63% enzyme
activity after 1 h incubation at 60益; PMSF could efficiently inhibit its activity, which confirmed it was a serine-type
carboxypeptidase. The bitterness of SPI and casein hydrolysates catalyzed by Alcalase significantly reduced after
treated with rOcpO and the degree of hydrolysis (DH) efficiently improved at the same time. |
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