State Key Laboratory of Food Science and Resources, Nanchang University, Nanchang, Jiangxi 330047, China.
National R&D Center for conventional Freshwater Fish Processing, Jiangxi Normal University, Nanchang 330022, China; Engineering Research Center of Freshwater Fish High-value Utilization of Jiangxi Province, Jiangxi Normal University, Nanchang, Jiangxi 330022, China.
Int J Biol Macromol. 2024 Nov;279(Pt 4):135469. doi: 10.1016/j.ijbiomac.2024.135469. Epub 2024 Sep 7.
In this study, the purpose was to screen novel angiotensin converting enzyme inhibitory peptides (ACEIPs) from tuna muscle taking two-steps enzymatic hydrolysis (Neutrase and Alkaline). Following isolation and purification by ultrafiltration, the Sephadex G-15 gel chromatography and reversed-phase high-performance liquid chromatography based on active-guide, the amino acid sequence was identified using Q-Orbitrap-MS/MS. Five peptides were chose synthesized based on the in silico screening methods. Among these, the two novel ACEIPs LTGCP and YPKP showed better inhibitory ability, and their corresponding IC values were 64.3 μM and 139.6 μM. Subsequently, the interaction mechanism of the best active peptide (LTGCP) against ACE was investigated by inhibitory pattern, molecular docking and molecular dynamic simulation. The result displayed that LTGCP was a mix-type inhibitor against ACE from the Lineweaver-Burk plots. LTGCP formed seven hydrogen bonds based on the molecular docking and the binding energy was -7.29 kcal/mol. LTGCP formed a stability complex with ACE based on the molecular dynamic simulation. Besides, LTGCP exhibited good stability in various temperature, pH and gastrointestinal digestion. Finally, the 0.125 mM ∼ 1.0 mM LTGCP exhibited no-toxic for Caco-2 cell. In summary, these findings showed that tuna was a good material to prepare ACEIPs and LTGCP may be the good potential antihypertensive drug or nutraceuticals.
在这项研究中，目的是通过两步酶解（中性蛋白酶和碱性蛋白酶）从金枪鱼肌肉中筛选新型血管紧张素转化酶抑制肽（ACEIP）。经超滤法分离和纯化后，采用基于活性导向的 Sephadex G-15 凝胶色谱法和反相高效液相色谱法，使用 Q-Orbitrap-MS/MS 鉴定氨基酸序列。根据计算机筛选方法选择合成了五种肽。其中，两个新型 ACEIP LTGCP 和 YPKP 表现出更好的抑制能力，其相应的 IC 值分别为 64.3 μM 和 139.6 μM。随后，通过抑制模式、分子对接和分子动力学模拟研究了最佳活性肽（LTGCP）与 ACE 的相互作用机制。结果表明，LTGCP 是 ACE 的混合型抑制剂，从 Lineweaver-Burk 图可以看出。LTGCP 基于分子对接形成了七个氢键，结合能为-7.29 kcal/mol。基于分子动力学模拟，LTGCP 与 ACE 形成了稳定的复合物。此外，LTGCP 在各种温度、pH 值和胃肠道消化条件下均表现出良好的稳定性。最后，0.125 mM～1.0 mM 的 LTGCP 对 Caco-2 细胞无毒性。总之，这些发现表明金枪鱼是制备 ACEIP 的良好材料，LTGCP 可能是一种具有良好潜力的抗高血压药物或营养保健品。
