Distirbution, identification and amino acid composition analysis of proteins in the secondary precipitate of soy sauce
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摘要: 以酱油上清液中可溶性蛋白质为对照,以静置-真空冷冻干燥法制备酱油二次沉淀,采用N-三（羟甲基）甘氨酸-十二烷基磺酸钠-聚丙烯酰胺凝胶电泳（Tricine-SDS-PAGE）、基质辅助激光解析电离飞行时间串联质谱（MALDI-TOF/TOF MS）和高效液相色谱（HPLC）法对比研究了酱油二次沉淀蛋白质的分子量分布、氨基酸组成特征和蛋白质的疏水性,并对酱油二次沉淀蛋白质进行了分子鉴定。结果表明:酱油二次沉淀蛋白质由分子量约为22.7ku（80.20%）和34.1ku（19.80%）的蛋白质组成,质谱鉴定结果证实前者为大豆球蛋白（Glycinin）G4亚基碱性端B3,后者为大豆球蛋白（Glycinin）G1酸性端A1a。氨基酸组成分析表明酱油二次沉淀蛋白质与酱油上清蛋白质氨基酸组成存在一定差异,酱油二次沉淀蛋白质的疏水性明显高于酱油上清蛋白质。本研究进一步深化了对酱油二次沉淀蛋白质的认识,为酱油二次沉淀问题的解决提供了新的依据。Abstract: In this paper, proteins in the supernatant of soy sauce were used as control, and the secondary precipitate of soy sauce were prepared by static settlement-lyophilization.The molecular weight distribution, amino acid composition and average hydrophobicity of proteins in the secondary precipitate of soy sauce were investigated by N-Tris[Hydroxymethyl]Methylglycine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis ( Tricine- SDS- PAGE) and high- performance liquid chromatography ( HPLC) , then proteins in the secondary precipitate of soy sauce were identified by matrix- assisted laser desorption ionisation- time of flight /time of flight mass spectrometry ( MALDI- TOF/TOF MS) . Results showed that proteins with molecular weights of approximate22.7ku ( 80.20%, the basic polypeptide B3of soy glycinin G4) and 34.1ku ( 19.80%, the acidic polypeptide A1a of soy glycinin G1) were identified as the majority of proteins in the secondary precipitate of soy sauce. Furthermore, remarkable difference in amino acid compostion was observed between proteins in the supernant and the secondary precipitate of soy sauce, average hydrophobicity analysis showed that average hydrophobicity of proteins in the secondary precipitate was significantly higher than that in proteins of the supernatant. This work could contribute to the systematical knowledge of proteins in the secondary precipitate of soy sauce, and shed some light on its solving way.
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Keywords:
- soy sauce /
- secondary precipitate /
- protein identification /
- amino acid composition
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