Quantitative structure-activity relationship studies of acteylcholinesterase-inhibitory peptides

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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.