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: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=689557
• 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.

生物活性多肽是存在于食物蛋白质线性链中的氨基酸的短序列(&lt；20个残基)，可以通过仔细的酶消化来释放。释放后，这些多肽成为潜在的强大试剂，可以恢复正常的生理过程。科学文献已经证实，氨基酸的线性排列是多肽活性类型和效力的重要决定因素。因此，这项建议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排列。人类AChE是一种酯酶，可以水解乙酰胆碱，乙酰胆碱是神经系统中的主要神经递质。AChE的过度活动与神经系统疾病有关，特别是痴呆症和阿尔茨海默病。然而，关于氨基酸类型和多肽链上的位置与抑制AChE活性之间的关系的信息很少。我的NSERC资助的Discovery研究项目的初步工作已经证明了使用酶消化从大麻种子蛋白中释放AChE抑制多肽的可行性。这项提议寻求继续胃酶的工作，以包括其他食物蛋白质(豌豆、油菜籽、亚麻籽、大豆、鹰嘴豆)，并最终导致发现新的AChE抑制多肽。因此，本方案的主要目的是分离、纯化和测定存在于这些植物蛋白的酶解物中的AChE抑制多肽的氨基酸序列。靶肽是那些含有10个或更少氨基酸的多肽，因为它们的小尺寸增加了穿越血脑屏障(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，水解物将通过1 kDa的超滤膜进行过滤，并收集流过的(渗透液)用于肽的纯化。纯化的多肽将经过高效液相色谱/串联质谱仪鉴定氨基酸序列，用于建立偏最小二乘回归分析的数据库。PLS模型将用于进一步发现新的AChE抑制多肽。利用Wistar大鼠模型，将评估具有最强抗乙酰胆碱酯酶效力的多肽通过血脑屏障和降低AChE大脑活动的能力。这项拟议的工作意义重大，因为这些结果将为AChE抑制多肽的氨基酸结构需求提供基本的科学信息。这种结构信息将使基于多肽的营养工具的开发成为可能，这些工具可以使与神经系统相关的生理障碍正常化。这项为期5年的拟议工作将培养3名博士后研究员、2名博士生、3名硕士研究生和5名本科生暑期研究学生。