基于SDS-PAGE对蜂蜜中蛋白质的研究

Analysis on protein in honey by SDS-PAGE

采用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)对蜂蜜中的蛋白进行分离,通过单一变量试验得出较优的电泳参数为:蛋白提取方法采用硫酸-钨酸钠法、上样量为20μg、分离胶浓度为12%。通过优化后的电泳条件对7种不同蜜源的天然蜂蜜和市售蜂蜜进行分离。结果分析得到天然蜂蜜中的蛋白质分子质量主要集中在(20~80)ku范围内,蛋白条带在(6~12)条之间,且不同蜜源蜂蜜中的蛋白条带具有很大的相似性;而市售蜂蜜的条带数仅为(4~6)条,蛋白含量与同蜜源天然蜂蜜相比明显减少。最后,将天然蜂蜜中的主要蛋白条带利用基质辅助激光解吸电离飞行时间二级质谱(MALDI-TOF/TOF MS)进行了鉴定,结果显示大部分蛋白为来源于意大利蜜蜂分泌的MRJPs(Majoy Royal Jelly Proteins)家族蛋白。

In this paper, proteins in honeys were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis（SDS-PAGE）. The optimum experimental parameters obtained through single variable test were： the protein extraction method was sulfuric acid-sodium tungstate method, the loading amount was 20 pg, and gel concentration was 12%. Proteins in seven different nectar sources of natural honey and commercial honey were separated through the optimized electrophoresis conditions. The result showed that the molecular weight of proteins from natural samples were mainly distributed between （20-80）ku, and the numbers of protein band were about 6-12. Besides, the proteins had great similarities in different sources of honey. The numbers of protein band in commercial honey were only 4-6, and the content of proteins decreased significantly compared with same nectar of natural honey. In addition, the major proteins in natural honey were identified by matrix-assisted laser desorption/ionization time-of-flight/time- of-flight mass spectrometry（MALDI-TOF/TOF MS）. The results showed that most of the proteins in honey were secreted by Apis mellifera called Majoy royal jelly proteins （MRJPs）.