犹他游动放线菌SW1311中阿库来菌素A酰基酶在棘白菌素类抗真菌药物合成中的应用（英文）
发布时间：2018-11-24 21:28
【摘要】：棘白菌素类抗真菌药物米卡芬净和阿尼芬净的合成工艺中包括一个关键步骤:水解除去FR901379分子和棘白菌素B(echinocandin B,ECB)分子的脂肪酸侧链,形成环状的6元多肽核心。FR901379和ECB的水解可以分别被FR901379酰基酶和阿库来菌素A酰基酶(aculeacin A acylase,AAC)催化,因此,FR901379酰基酶和AAC的发掘、表征和生产在米卡芬净和阿尼芬净的工业生产上具有重要的应用价值。本研究首先筛选到了犹他游动放线菌SW1311,发现该菌株发酵液具有酰基酶活性,并摸索了不同发酵条件对酰基酶活性的影响。然后将犹他游动放线菌SW1311中的AAC基因克隆到改造过的质粒载体p IJ8660中,并将该质粒转化到天蓝色链霉菌(Streptomyces coelicolor)A3(2)中对AAC基因进行高表达,得到重组菌株sSCO-AAC。最后将sSCO-AAC生产的AAC活性和所需培养时间与犹他游动放线菌SW1311进行比较,表征了sSCO-AAC的发酵液水解FR901379的反应。结果表明,犹他游动放线菌SW1311中的酰基酶具有水解FR901379和青霉素V的酰基活性。用重组菌株sSCO-AAC生产的AAC活性比犹他游动放线菌SW1311的高4.6倍,且该重组菌株所需的培养时间比犹他游动放线菌SW1311缩短了30%。该结果不仅将ACC的应用范围从阿尼芬净合成拓宽到了米卡芬净合成,而且还揭示出天蓝色链霉菌A3(2)可以作为一个良好的AAC表达菌株。本研究对阿尼芬净和米卡芬净的工业化生产具有潜在的应用价值。
[Abstract]:One of the key steps in the synthesis of mikafenem and Amifen is the hydrolysis of fatty acid side chains of FR901379 molecules and B (echinocandin FR901379 molecules. The hydrolysis of FR901379 and ECB can be catalyzed by FR901379 acylase and acurylin A acylase (aculeacin A acylase,AAC, respectively. Therefore, the discovery of FR901379 acylase and AAC, Characterization and production have important application value in the industrial production of mikafennet and arnifen. In this study, first of all, we found that the fermentation broth of SW1311, had acylase activity, and explored the effect of different fermentation conditions on the activity of acylase. Then, the AAC gene from SW1311 was cloned into the modified plasmid vector p IJ8660, and the AAC gene was overexpressed in Streptomyces aquilinum (Streptomyces coelicolor) A3 (2). The recombinant strain sSCO-AAC. was obtained. Finally, the activity and culture time of AAC produced by sSCO-AAC were compared with that of SW1311, and the reaction of hydrolyzing FR901379 in sSCO-AAC fermentation broth was characterized. The results showed that the acylase in SW1311 had the activity of hydrolyzing FR901379 and penicillin V. The AAC activity of the recombinant strain sSCO-AAC was 4. 6 times higher than that of SW1311, and the culture time of the recombinant strain was 30 times shorter than that of SW1311. This result not only broadens the application range of ACC from ANFN synthesis to mikafennet synthesis, but also reveals that Streptomyces aquilinus A3 (2) can be used as a good AAC expression strain. This study has potential application value for the industrial production of arnifen and mikafennet.
【作者单位】： 华东医药集团新药研究院有限公司;浙江大学求是学院;浙江大学生命科学学院;
【基金】：Projects supported by Zhejiang Provincial Natural Science Foundation of China(No.LR16H300001) National Natural Science Foundation of China(No.31670008)
【分类号】：R914
，
本文编号：2355125
[Abstract]:One of the key steps in the synthesis of mikafenem and Amifen is the hydrolysis of fatty acid side chains of FR901379 molecules and B (echinocandin FR901379 molecules. The hydrolysis of FR901379 and ECB can be catalyzed by FR901379 acylase and acurylin A acylase (aculeacin A acylase,AAC, respectively. Therefore, the discovery of FR901379 acylase and AAC, Characterization and production have important application value in the industrial production of mikafennet and arnifen. In this study, first of all, we found that the fermentation broth of SW1311, had acylase activity, and explored the effect of different fermentation conditions on the activity of acylase. Then, the AAC gene from SW1311 was cloned into the modified plasmid vector p IJ8660, and the AAC gene was overexpressed in Streptomyces aquilinum (Streptomyces coelicolor) A3 (2). The recombinant strain sSCO-AAC. was obtained. Finally, the activity and culture time of AAC produced by sSCO-AAC were compared with that of SW1311, and the reaction of hydrolyzing FR901379 in sSCO-AAC fermentation broth was characterized. The results showed that the acylase in SW1311 had the activity of hydrolyzing FR901379 and penicillin V. The AAC activity of the recombinant strain sSCO-AAC was 4. 6 times higher than that of SW1311, and the culture time of the recombinant strain was 30 times shorter than that of SW1311. This result not only broadens the application range of ACC from ANFN synthesis to mikafennet synthesis, but also reveals that Streptomyces aquilinus A3 (2) can be used as a good AAC expression strain. This study has potential application value for the industrial production of arnifen and mikafennet.
【作者单位】： 华东医药集团新药研究院有限公司;浙江大学求是学院;浙江大学生命科学学院;
【基金】：Projects supported by Zhejiang Provincial Natural Science Foundation of China(No.LR16H300001) National Natural Science Foundation of China(No.31670008)
【分类号】：R914
，
本文编号：2355125
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