Quantitative structure-activity relationship studies of acteylcholinesterase-inhibitory peptides

乙酰胆碱酯酶抑制肽的定量构效关系研究

• 批准号: RGPIN-2018-06019
• 负责人: Aluko, Rotimi
• 金额: $ 3.42万
• 依托单位: University of Manitoba
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=714036
• 关键词: Quantitative; structure; activity; relationship; studies

Bioactive peptides are short sequences (<20 residues) of amino acids present within the linear chain of food proteins and can be released through careful enzymatic digestion. Upon release, these peptides become potentially powerful agents that can restore normal physiological processes. It has been established in scientific literature that the linear arrangement of amino acids is an important determinant of the type and potency of activity of the peptide. Therefore, the focus of this proposal is to determine the amino acid positional arrangements that influence potency of acetylcholinesterase (AChE)-inhibitory peptides. Human AChE is an esterase that hydrolyzes acetylcholine, the main neurotransmitter in the nervous system. Excessive activities of AChE have been implicated in disorders of the nervous system, especially dementia and Alzheimer's disease. However, there is scant information on the relationships of amino acid type and position on the peptide chain with ability to inhibit AChE activity. Preliminary work from my NSERC-funded Discovery research program has demonstrated the feasibility of using enzyme digestion to release AChE-inhibitory peptides from hemp seed proteins. This proposal seeks to continue the pepsin work to include other other food proteins (peas, canola, flaxseed, soybean, chickpea) and ultimately lead to the discovery of novel AChE-inhibitory peptides. Therefore, the main objectives of this proposal are to fractionate, purify, and determine the amino sequence of AChE-inhibitory peptides present in the enzymatic hydrolysates of these plant proteins. Target peptides are those with 10 or less amino acids because their small sizes increase the potential to cross the blood-brain barrier (BBB), enter the brain, bind to AChE and reduce excessive breakdown of acetylcholine. Thus, the hydrolysates will be filtered through a 1 kDa ultrafiltration membrane and the flow-through (permeate) collected and used for peptide purification. Purified peptides will be subjected to liquid chromatography/tandem mass spectrometry to identify amino acid sequences, which will be used to build databases for partial least square (PLS) regression analysis. PLS models will be used for additional discovery of new AChE-inhibitory peptides. Using the Wistar rat model, peptides with strongest potency against AChE will be evaluated for ability to cross the BBB and reduce brain activity of AChE. The proposed work is significant because the results will provide fundamental scientific information on the amino acid structural requirements for AChE-inhibitory peptides. Such structural information will enable development of peptide-based nutritional tools that can normalize physiological disorders associated with the nervous system. The proposed work for this 5-yr cycle will train 3 postdoctoral fellows, 2 PhD students, 3 MSc students and 5 undergraduate summer research students.

生物活性肽是存在于食物蛋白质线性链中的短序列（<20个残基）氨基酸，可以通过仔细的酶消化释放。一旦释放，这些肽成为潜在的强大的代理商，可以恢复正常的生理过程。在科学文献中已经确定，氨基酸的线性排列是肽的活性类型和效力的重要决定因素。因此，本提案的重点是确定影响乙酰胆碱酯酶（AChE）抑制肽效力的氨基酸位置安排。人类乙酰胆碱酯酶是一种水解乙酰胆碱的酯酶，乙酰胆碱是神经系统中的主要神经递质。乙酰胆碱酯酶的过度活动与神经系统疾病，特别是痴呆和阿尔茨海默病有关。然而，有很少的信息的氨基酸类型和位置的肽链上的能力，抑制乙酰胆碱酯酶活性的关系。我的NSERC资助的探索研究计划的初步工作已经证明了使用酶消化从大麻籽蛋白中释放乙酰胆碱酯酶抑制肽的可行性。这项提案旨在继续胃蛋白酶的工作，包括其他食物蛋白质（豌豆，油菜籽，亚麻籽，大豆，鹰嘴豆），并最终导致新的乙酰胆碱酯酶抑制肽的发现。因此，本提案的主要目标是对这些植物蛋白的酶水解产物中存在的乙酰胆碱酯酶抑制肽进行分离、纯化和确定其氨基酸序列。靶肽是具有10个或更少氨基酸的那些肽，因为它们的小尺寸增加了穿过血脑屏障（BBB）、进入大脑、与AChE结合并减少乙酰胆碱过度分解的可能性。因此，水解产物将通过1 kDa超滤膜过滤，收集流出物（渗透物）并用于肽纯化。纯化的肽将进行液相色谱/串联质谱法以鉴定氨基酸序列，其将用于构建偏最小二乘（PLS）回归分析的数据库。PLS模型将用于新的乙酰胆碱酯酶抑制肽的额外发现。使用Wistar大鼠模型，将评估对AChE具有最强效力的肽穿过BBB和降低AChE大脑活动的能力。拟议的工作是有意义的，因为结果将提供基本的科学信息的氨基酸结构要求的乙酰胆碱酯酶抑制肽。这种结构信息将使基于肽的营养工具的开发成为可能，这些营养工具可以使与神经系统相关的生理障碍正常化。这个5年周期的拟议工作将培养3名博士后研究员，2名博士生，3名硕士生和5名本科夏季研究生。