Novel Chymotrypsin Purified and Biochemically Characterized from Digestive Organs of Bigfin Reef Squid (Sepioteuthis lessoniana)

IF 3.6 Q2 ENVIRONMENTAL SCIENCES

Resources Pub Date : 2024-05-14 DOI:10.3390/resources13050067

Jirapan Satjarak, S. Klomklao, Yi Zhang, K. Thongprajukaew

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Abstract

Chymotrypsin from the digestive organs of bigfin reef squid, a major commercial squid species in Thailand, was purified to 41-fold with 5.7% yield by a sequential purification process including ammonium sulfate precipitation, size exclusion, and ion exchange chromatography. According to SDS-PAGE, the molecular weight of purified chymotrypsin was 43 kDa. Native-PAGE analysis revealed a single band for this purified enzyme. The optimum pH and temperature for chymotrypsin activity of the purified enzyme were a pH of 7.0 with a temperature of 55 °C. The purified chymotrypsin remained stable throughout a wide range of pH levels (6–11) and at relatively high temperature (55 °C). It was significantly inhibited by PMSF and TPCK. The values of the kinetic constants Km and Kcat were found to be 1.33 mM and 31.46 s−1, respectively. The purified chymotrypsin has the N-terminal amino acid IVGGQEATPGEWPWQAALQV. This study provided new information about the biochemical properties of pure chymotrypsin from bigfin reef squid, which will be useful in the future investigation, aquaculture, and application of bigfin reef squid.

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从大鳍礁鱿鱼（Sepioteuthis lessoniana）消化器官中纯化出的新型糜蛋白酶及其生物化学特征

通过硫酸铵沉淀、尺寸排阻和离子交换色谱等连续纯化过程，从泰国一种主要商业鱿鱼品种--大鳍礁鱿鱼的消化器官中纯化了 41 倍的糜蛋白酶，产率为 5.7%。根据 SDS-PAGE 分析，纯化的糜蛋白酶分子量为 43 kDa。原生聚合酶链分析显示，纯化的糜蛋白酶只有一个条带。纯化酶的糜蛋白酶活性的最佳 pH 值和温度为 pH 值 7.0 和温度 55 ℃。纯化的糜蛋白酶在很宽的 pH 值范围（6-11）和相对较高的温度（55 °C）下都保持稳定。它受到 PMSF 和 TPCK 的明显抑制。动力学常数 Km 和 Kcat 的值分别为 1.33 mM 和 31.46 s-1。纯化的糜蛋白酶的 N 端氨基酸为 IVGGQEATPGEWPWQAALQV。该研究为纯化的大翅礁鱿鱼糜蛋白酶的生化特性提供了新的信息，将有助于今后大翅礁鱿鱼的研究、养殖和应用。

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来源期刊

Resources Environmental Science-Nature and Landscape Conservation

CiteScore

7.20

自引率

6.10%

发文量

审稿时长

11 weeks

期刊介绍： Resources (ISSN 2079-9276) is an international, scholarly open access journal on the topic of natural resources. It publishes reviews, regular research papers, communications and short notes, and there is no restriction on the length of the papers. Our aim is to encourage scientists to publish their experimental and theoretical research in as much detail as possible. Full experimental and methodical details must be provided so that the results can be reproduced. There are, in addition, unique features of this journal: manuscripts regarding research proposals and research ideas will be particularly welcomed, electronic files or software regarding the full details of the calculation and experimental procedure, if unable to be published in a normal way, can be deposited as supplementary material. Subject Areas: natural resources, water resources, mineral resources, energy resources, land resources, plant and animal resources, genetic resources, ecology resources, resource management and policy, resources conservation and recycling.