Abstract：
Cold-adapted proteases are capable of efficient protein hydrolysis at reduced temperatures, which offer significant potential applications in the area of low temperature food processing. In this paper, we attempted to characterize cold-adapted proteases from Antarctic krill. Antarctic krill possesses an extremely active autolytic enzyme system in their bodies, and the production of peptides and free amino acids accompanies the rapid breakdown of muscle proteins following the death. The crucial role of trypsin in this process is recognized. A cold-adapted trypsin named OUC-Pp-20 from Antarctic krill genome was cloned and expressed in Pichia pastoris. Recombinant trypsin is a monomeric protein of 26.8 ± 1.0 kDa with optimum reaction temperature at 25 °C. In addition, the catalytic specificity of OUC-Pp-20 was assessed by identifying its hydrolysis sites through LC-MS/MS. OUC-Pp-20 appeared to prefer Gln and Asn at the P1 position, which is an amino acid with an amide group in its side chain. Hydrolysis reactions on milk and shrimp meat revealed that it can effectively degrade allergenic components in milk and arginine kinase in shrimp meat. These findings update the current knowledge of cold-adapted trypsin and demonstrate the potential application of OUC-Pp-20 in low temperature food processing.
摘要：
冷适应蛋白酶能够在降低的温度下有效的蛋白质水解，在低温食品加工领域提供了重要的潜在应用。在本文中,我们试图表征来自南极磷虾的冷适应蛋白酶。南极磷虾体内具有极为活跃的自溶酶系统,肽和游离氨基酸的产生伴随着死亡后肌肉蛋白的快速分解。胰蛋白酶在这一过程中的关键作用是公认的。克隆了来自南极磷虾基因组的冷适应胰蛋白酶OUC-Pp-20，并在巴斯德毕赤酵母中表达。重组胰蛋白酶是26.8±1.0kDa的单体蛋白，最佳反应温度为25℃。此外,通过LC-MS/MS鉴定其水解位点来评估OUC-Pp-20的催化特异性。OUC-Pp-20似乎在P1位置更喜欢Gln和Asn，它是在其侧链上带有酰胺基的氨基酸。对牛奶和虾肉的水解反应表明，它可以有效地降解牛奶中的致敏成分和虾肉中的精氨酸激酶。这些发现更新了冷适应胰蛋白酶的最新知识，并证明了OUC-Pp-20在低温食品加工中的潜在应用。
