Screening,Identification And Function Of Immunomodulating Peptide Receptor From Human Ovarian Carcinoma Cells

Objective To screen, isolate and identify the receptor of immunomodulating peptide(PGPIPN) in ovarian cancer cell lines (SKOV3).Methods SKOV3and PGPIPN labeled fluorescein FITC were incubation together,PGPIPN binding sites at the cell were observed under confocal laser scanningmicroscope. The whole membrane proteins were extracted from ovarian cancer cell bymembrane protein extraction kit. Taking PGPIPN as affinity chromatography ligand,the receptors were purified by NHS-activated affinity chromatography, which wereenriched by vacuum freeze drying apparatus. The screening result was preliminaryidentificated by SDS-PAGE and HPLC-ESI-MS. The protein bands that specificallybound to PGPIPN were screened, and then identified and analyzed by electrosprayionization mass spectrometry and bioinformatics. The characteristics of the receptorsto bind PGPIPN was proved by the fluorescence obversation of the fluorescencelabeling peptide. A certain concentration of the PGPIPN, some differentconcentrations of receptors and ovarian cancer cells were togather incubation withMTT, to observe the influence of the proliferation/apoptosis in SKOV3.Results Compared with the control group, the fluorescence labeled peptides wereobserved to locate in the ovarian cancer cell membrane by the confocal laser scanningmicroscope.The membrane proteins were successful extracted by using reagent kit.Receptor has been screened and purified from membrane proteins by using affinitychromatography. The receptors were primary identified by HPLC-ESI-MS, andcharacteristics of the receptor bind PGPIPN was also proved by the fluorescenceobversation of the fluorescence labeling peptide. The receptor function was primary identified by the study on proliferation of ovarian cancer cells, the results were ATPsynthase,basigin,et al.Conclusion PGPIPN receptor locates at the cell membrane. The anticancer of PGPIPN,which can inhibit proliferation and induce apoptosis in ovarian carcinoma cell linesSKOV3,may effect through binding membrane protein to trigger signal transduction.This study has laid the foundation for further probing the anticancer mechanism ofPGPIPN.