还原性青刺果果实蛋白生物化学特性的研究

摘要

分离还原性青刺果果实蛋白,研究其生物化学特性.使用改良的铁氰化钾还原法评价青刺果果实蛋白的还原能力,酸溶醇沉法提取青刺果果实蛋白.基于青刺果果实蛋白PSP4最强的还原能力,通过SDS-PAGE、氨基酸自动分析、红外光谱分析、圆二色光谱分析、差示扫描量热分析,测定PSP4的氨基酸构成、纯度及分子大小、二级结构和变性温度等生物化学特性.结果显示75％乙醇沉淀青刺果果实蛋白PSP4还原力半数有效浓度为24.7 mg/mL,SDS-PAGE电泳呈现清晰的3条谱带,相对分子质量分别为11 KDa、14 KDa和34 KDa,热变性温度为69.83℃,还原性青刺果果实蛋白PSP4具有典型的酰胺Ⅰ(1690～1650 cma)、酰胺Ⅱ(1610 ～ 1370cm-1)、酰胺Ⅲ(1100～1030 cm-1)、酰胺A(3400～3230 cm-1)和酰胺B(3000～ 2800 cm-1)的吸收谱带,二级结构包含有α-螺旋(5.2％)、β-折叠(43.1％)和无规卷曲(27.6％).研究表明,以PSP4为代表的还原性青刺果果实蛋白由多条肽链构成、β-折叠占优势、较高的热变性温度是其重要生物化学特性,具有作为生物功能物质的优良特征.%Reducing proteins were separated from Prinsepiautilis Roylefmit and the excellent biochemical properties were investigated.Improved reducing power evaluation method with Potassium Ferricyanide was used.P.utilis fruit proteins were extracted and purified by acid dissolution and ethanol precipitation.Because of the strongest reducing power of P.utilisfruit special protein 4,SDS-PAGE,amino acids automatic analysis,infrared spectroscopy,circular dichroism spectroscopy and differential scanning calorimeter were applied to determine composition,purity,molecular size,secondary structure,denaturation temperature and other biochemical characteristics of amino acids in P.utilisfruit special protein 4,respectively.The reducing power of P.utilisfruit special protein 4 precipitated by 75％ ethanol was observed.Its half effective concentration of reducing power was 24.7 mg/mL.The extracted P.utilisfruit special protein 4 presented as three distinct bands by SDS-PAGE,and the relative molecular mass were 11,14 and 34 KDa respectively.The denatured temperature of P.utilisfruit special protein 4 was 69.83 ℃.The secondary structure ofP.utilisfruit special protein 4 was primarily presented asβ-sheet (43.1％) and the random structure (27.6％),whereas alpha-helix (5.2％) occupies the minority.Research showed that the representative of P.utilisfruit special protein 4 was composed of several peptide chains,β-sheet in a dominant position,higher thermal denaturation temperature.These were important biochemical characteristics.P.utilisfruit special protein 4 with reducing capability was revealed as potential functional substances with excellent characteristics.