Screening And Identification Of Mouse Proteins Interacted With NLRP2

Mouse oocytes,early embryos and ovaries were studied in this experiment.Proteins interacted with NLRP2 were screened and identified by employing a series of techniques,including bioinformatics,western blot,immunofluorescence,RNA interference,co-immunoprecipitation,and mass spectrography analysis.The results are as follows:The structure and physicochemical property of Nlrp2 gene and NLRP2 protein were analyzed by bioinformatics.The result showed that Nlrp2 gene is localized at chromosome 7A1 and NLRP2 protein contained 1046 amino acid,its molecular weight is 118641.0,the isoelectric charge is 6.27 and the molecular formula is C5286H8453N1387O1554S73.NLRP2 proteins showed no signal sequence and transmembrane structure.However,there are three domains and two P-loop structures in NLRP2 protein.FAF1,MEFV and NAIP1 were predicted to interact with NLRP2 protein,respectively.To determine if FAF1,MEFV and NAIP1 interact with NLRP2 protein,reciprocal experiments were carried out using co-immunoprecipitation and western blot.The result showed that there are no interactions between NLRP2 protein with MEFV or NAIP1,while NLRP2 interact with FAF1 protein.Furthermore,NLRP2 and FAF1 protein have identical co-localization in mouse oocytes and early embryos by immunofluorescence and confocal microscopy.Moreover,Knockdown of Nlrp2 or Fafl in zygotes led to the sharply decline of its interacted protein.MYH11 and RS3 protein,which may be interact with NLRP2 protein,were screened by co-immunoprecipitation,SDS-PAGE and mass spectrography analysis.Reciprocal experiments showed that MYH11 and RS3 protein are clearly detectable in anti-NLRP2 immunoprecipitates,meanwhile,NLRP2 protein is clearly detectable in anti-MYH11 or anti-RS3 immunoprecipitates.Further experiments should be carried out to verify the interaction between NLRP2 and MYH11 or RS3 protein and uncovered its physiological function in early embryo development in the mouse.