应用表面增强激光解吸电离飞行时间质谱技术筛选牙龈卟啉单胞菌共同外膜蛋白

Analysis of outer membrane proteins in various strains of Porphyroronas gingivalis by surface enhanced laser desorption/ionization time-of-flight mass spectrometry

目的 应用表面增强激光解吸电离飞行时间质谱（surface enhanced laser desorption/ionization time-of-flight mass spectrometry,SELDI-TOF-MS）技术筛选多种牙龈卟啉单胞菌（Porphyromonas gingivalis,Pg）共同外膜蛋白（outer membrane protein,OMP）,为针对多种Pg设计具有交叉保护作用的疫苗提供候选靶抗原.方法 应用超速离心法提取PgATCC33277、Pgw83、Pg301和Pg381菌株外膜蛋白,SELDI疏水性蛋白芯片H50检测OMP,Biomarker Wizard软件分析4种菌株OMP质谱图.结果 OMP质谱图显示,PgATCC33277、PgW83、Pg301和Pg381分别有71、74、76和72个蛋白质峰,并存在13种共同OMP 在美国国立生物信息中心蛋白库中鉴定到一种已知蛋白gil116636495,其功能尚不清楚.结论 发现13种共同OMP可作为牙周病疫苗候选抗原,证实基于SELDI-TOF-MS技术适合检测小相对分子质量（＜20000）、低丰度、疏水性蛋白,且操作简单、重复性好,可用于大规模寻找Pg的共同OMP.

Objective To analyze the common outer membrane proteins （OMP）from Porphyromonas gingivalis （Pg）by surface enhanced laser desorption/ionization time-of-flight mass spectrometry （SELDI-TOF-MS） and to provide antigens for the subsequent experiments in screening vaccine for periodontitis therapy. Methods Four strains of Pg were cultured under anaerobic conditions. The common OMP was extracted through ultracentrifugation and SELDI-TOF-MS was employed to detect the expressions of proteomes by chip H50. The data was analyzed by Biomarker Wizard. ResultsFour kinds of strains of OMP fingerprint spectrum were obtained. Seventy-one proteins of PgATCC33277, 74 proteins of PgW83, 76 proteins of PgW301 and 72 proteins of Pg381 were captured by chip H50. Thirteen common proteins were identified according to fingerprint spectrum. There was only 1 of the 13 common proteins identified in NCBI protein bank. ConclusionsSELDI-TOF-MS has good reproducibility and high sensibility and can be used to identify the common OMP of Pg. The 13 proteins have a potential value in the screening vaccine candidate antigen sites for periodontitis.