十肽对变异链球菌作用的差异蛋白质组学研究

摘要

目的：  利用差异蛋白质组学技术检测十肽对变异链球菌蛋白的作用反应，初步鉴定并验证变异链球菌致龋过程中的部分重要蛋白质。方法  人工合成十肽（氨基酸序列：KKVVFKVKFK-NH2），利用双向电泳技术分离十肽作用前后变异链球菌的总蛋白，采用质谱分析联合生物信息学技术鉴定十肽作用前后发生差异性改变的变异链球菌蛋白质，并在蛋白质水平验证关键差异蛋白质烯醇酶的表达情况。结果  十肽作用前后变异链球菌蛋白质发生明显变化，其中发生差异性改变的蛋白质功能主要包括降解碳水化合物（经糖酵解途径），蛋白质折叠、结合、运输及蛋白质翻译等。Western blot验证结果显示，十肽处理后烯醇酶的表达量明显降低。结论  人工合成抗菌肽十肽处理前后变异链球菌的蛋白质表达发生明显变化，推测其可能通过改变烯醇酶等标志性蛋白的表达而达到防龋作用。%Objective  To compare the protein profiles between decapeptide-treated and untreated planktonic cells of Streptococcus mutans (S.mutans) by differential proteomic analysis to determine and identify the key proteins.Methods  In our previous study, we investigated decapeptide (KKVVFKVKFK–NH2), which was a novel adenosine monophosphate. Com-pared with other oral pathogens tested, decapeptide had a preferential antibacterial activity against S. mutans. It also inhibited S. mutans biofilm formation and reduced the one-day developed biofilm. In the present study, we first synthesized decapep-tide, and then compared the protein profiles between decapeptide-treated and untreated planktonic cells of S. mutans by two-dimensional gel electrophoresis and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. We also verified different expressions of key protein enolase in the protein level. Results  The results showed that decapeptide altered the protein expression of planktonic S. mutans. These proteins were functionally involved in carbohydrate degradation by gly-colysis, protein folding, conjunction, transport, translation, adenosine triphosphate binding, protein binding, sequence-specific DNA binding, transcription factor activity, and two-component response regulator activity. Western blot results showed that enolase protein expression decreased obviously in decapeptide-treated cells of S. mutans. Conclusion  The protein expression of S. mutans significantly changed after synthetic antimicrobial decapeptide treatment, suggesting that decapeptide may pre-sent a preferential effect on oral caries by changing the expression of certain key proteins, such as enolase protein.