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Aspergillus oryzae acetamidase catalyzes degradation of ethyl carbamate.
Journal of Bioscience and Bioengineering ( IF 2.8 ) Pub Date : 2020-08-26 , DOI: 10.1016/j.jbiosc.2020.07.015
Kazuo Masaki 1 , Kanako Fujihara 2 , Dararat Kakizono 3 , Taichi Mizukure 2 , Masaki Okuda 1 , Nobuhiko Mukai 3
Affiliation  

Urethanase (EC 3.5.1.75) catalyzes the hydrolysis of ethyl carbamate (EC) to ethanol, carbon dioxide, and ammonia. From our recent study, we expected that an acetamidase encoded by amdS of Aspergillus oryzae may catalyze the degradation of EC because it is homologous with a Candida parapsilosis urethanase (CPUTNase) recently identified. Urethanase is a prospective candidate to reduce EC in alcoholic beverages, but knowledge of this enzyme is very limited. Recombinant AmdS was expressed to study its enzymatic properties. Purified AmdS was identified as a homo-tetramer consisting of four 60 kDa units and exhibited urethanase activity. In a 20% ethanol solution, AmdS had 65% activity compared with a solution without ethanol. Residual activity after 18 h indicated that AmdS was stable in 0%–40% ethanol solutions. The optimum temperature of AmdS was 40 °C. This enzyme showed urethanase activity at pH 6.4–9.6 and exhibited its highest activity at pH 9.6. The Km value of AmdS for EC was 8.2 mM, similar to the Km value (7.6 mM) of CPUTNase. AmdS showed activity not only for EC and acetamide but also other amide compounds. In this study, we investigated the enzymatic properties of AmdS that was identified as acetamidase and showed that an amidase can be an enzymatic candidate that degrades EC.



中文翻译:

米曲霉乙酰酰胺酶催化氨基甲酸乙酯的降解。

脲酶(EC 3.5.1.75)催化氨基甲酸乙酯(EC)水解为乙醇,二氧化碳和氨。从我们最近的研究中,我们预计到编码乙酰胺的amdS米曲霉可以催化EC的下降,因为它是同源同一个近平滑念珠菌最近发现了尿烷酶(CPUTNase)。尿烷酶是减少酒精饮料中EC的潜在候选物,但是对该酶的了解非常有限。表达重组AmdS来研究其酶学性质。纯化的AmdS被鉴定为由四个60 kDa单位组成的同型四聚体,并显示出尿烷酶活性。与不含乙醇的溶液相比,在20%的乙醇溶液中,AmdS的活性为65%。18小时后的残留活性表明AmdS在0%–40%乙醇溶液中稳定。AmdS的最佳温度为40°C。该酶在pH 6.4-9.6时显示尿烷酶活性,在pH 9.6时显示其最高活性。EC的AmdS的K m值为8.2 mM,类似于K mCPUTNase的最大值(7.6 mM)。AmdS不仅对EC和乙酰胺都有活性,而且对其他酰胺化合物也有活性。在这项研究中,我们调查了被鉴定为乙酰酰胺酶的AmdS的酶促性质,并表明酰胺酶可以是降解EC的候选酶。

更新日期:2020-08-26
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