Abstract：
:Kumamolisin from Alicyclobacillus sendaiensis strain NTAP-1 is a serine protease with collagenase activity. After molecular engineering, a kumamolisin mutant, named Kuma030, was obtained with high proteolytic activity against gluten, which might cause celiac disease. Kuma030 exhibited its potential application in industrial and medicine, while challenges remained of its large-scale purification and production. In the studies here, we successfully overexpressed the Kuma030 in E. coli BL21 (DE3) by anchoring a SUMO (Small Ubiquitin-like Modifier) fusion protein at its N-terminal end. In addition, a fast protein purification procedure was developed according to the acidophilic and thermophilic properties of Alicyclobacillus sendaiensis. After a simple acid treatment followed by a heat treatment, a total of 9.9 mg functional Kuma030 was quickly obtained form 1 L LB media culture. This purified Kuma030 was confirmed to be functional to cleave the PQ sequences in a designed protein substrate, and the gluten in actual food samples, such as whole wheat bread and beer, in a fast manner. Our studies provided an efficient strategy for the overexpression and purification of functional Kuma030 in E. coli, which might expand its broad practical applications.
journal_name
Enzyme Microb Technoljournal_title
Enzyme and microbial technologyauthors
Liu H,Fan X,Song H,Hu X,Zhang G,Yu C,Yi Ldoi
10.1016/j.enzmictec.2019.05.013subject
Has Abstractpub_date
2019-10-01 00:00:00pages
109356eissn
0141-0229issn
1879-0909pii
S0141-0229(19)30085-7journal_volume
129pub_type
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journal_title：Enzyme and microbial technology
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journal_title：Enzyme and microbial technology
pub_type： 杂志文章
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journal_title：Enzyme and microbial technology
pub_type： 杂志文章
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journal_title：Enzyme and microbial technology
pub_type： 杂志文章
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pub_type： 杂志文章
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journal_title：Enzyme and microbial technology
pub_type： 杂志文章
doi：10.1016/j.enzmictec.2014.09.001
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journal_title：Enzyme and microbial technology
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journal_title：Enzyme and microbial technology
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journal_title：Enzyme and microbial technology
pub_type： 杂志文章
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journal_title：Enzyme and microbial technology
pub_type： 杂志文章
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更新日期：2018-11-01 00:00:00
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journal_title：Enzyme and microbial technology
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更新日期：2000-03-01 00:00:00
