Structure-function properties of food protein-derived calmodulin-binding peptides: in vivo studies

食物蛋白衍生的钙调蛋白结合肽的结构-功能特性：体内研究

• 批准号: 249890-2013
• 负责人: Aluko, Rotimi
• 金额: $ 4.15万
• 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: 2014
• 资助国家: 加拿大
• 起止时间: 2014-01-01 至 2015-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=547439
• 关键词: Structure; function; properties; food; protein

Calmodulin (CaM) is a 16.7 kDa multifunctional calcium-binding protein that is a proven major translator of intracellular calcium message. CaM is necessary for calcium-dependent control of many cellular events including cell proliferation, cell division and neurotransmission. Excessive activities of CaM-dependent enzymes can lead to metabolic disorders such as diabetes, hypertension, abnormal cell growth and arthritis. Therefore, peptides that bind to CaM and inhibit activity of CaM-dependent enzymes may be used to formulate nutritional supplements for preventing onset or progression of chronic ailments. My previous NSERC-funded work led to the discovery of several food (flaxseed and yellow field pea) protein-derived peptide sequences that work as in vitro inhibitors of CaM-dependent enzymes such as phosphodiesterase 1 (CaMPDE) and the nitric oxide synthases (NOSs). Therefore, the main objective of current proposal is to determine ability of these peptides (2-7 amino acid residues) to escape digestion in the gastrointestinal tract and be absorbed intact into the blood circulatory system. The work will also determine inhibitory ability of the absorbed peptides against tissue activities of CaMPDE and NOS enzymes (endothelial, neuronal and inducible), including peptide-induced reductions in levels of endogenous toxic free radicals as well as increased level of antioxidant capacity. To determine in vivo absorption properties, Wistar rats will be fed purified peptides or protein hydrolysate fractions with known peptide compositions. Peptide composition of the blood will be determined after gavage using liquid chromatography/mass spectrometry techniques to determine if absorption occurred. Oral feeding of peptides to rats subjected to ethanol- or lipopolysaccharide-induced oxidative stress will be followed by determination of plasma and organ tissue levels of toxic free radicals, especially superoxide whose production (catalyzed by endothelial NOS) becomes greatly enhanced during oxidative stress. The proposed work could enhance development of novel peptide-based supplements that can be used as preventive agents against accumulation of toxic free radicals in the body.

钙调蛋白(CaM)是一种16.7 kDa的多功能钙结合蛋白，是细胞内钙信息的主要转运体。钙调素是许多细胞活动的钙依赖调控所必需的，包括细胞增殖、细胞分裂和神经传递。CaM依赖的酶活性过高会导致代谢紊乱，如糖尿病、高血压、细胞异常生长和关节炎。因此，结合CaM并抑制CaM依赖酶活性的多肽可用于研制预防慢性疾病发生或进展的营养补充剂。我之前由NSERC资助的工作导致发现了几种食物(亚麻籽和黄豆)蛋白质衍生的多肽序列，它们在体外可以作为CaM依赖的酶的抑制物，如磷酸二酯酶1(CaMPDE)和一氧化氮合酶(NOSS)。因此，目前建议的主要目标是确定这些多肽(2-7个氨基酸残基)逃避胃肠道消化并被完整地吸收到血液循环系统的能力。这项工作还将确定被吸收的多肽对组织中CaMPDE和NOS酶(内皮、神经元和诱导性)活性的抑制能力，包括多肽诱导的内源性有毒自由基水平的降低以及抗氧化能力的提高。为了确定体内吸收特性，Wistar大鼠将喂食纯化的多肽或已知多肽组成的蛋白质水解物部分。血液中的多肽成分将在灌胃后使用液相色谱/质谱仪技术进行测定，以确定是否发生吸收。给乙醇或脂多糖诱导的氧化应激大鼠口服多肽后，将测定血浆和器官组织中有毒自由基的水平，特别是在氧化应激过程中产生的超氧化物歧化酶(由内皮型一氧化氮合酶催化)大大增加。这项拟议的工作可能会促进基于多肽的新型补充剂的开发，这些补充剂可以用作预防有毒自由基在体内积累的防腐剂。