鉴定免疫细胞中DNA疫苗结合蛋白

为研究DNA疫苗从细胞质到细胞核的过程,对免疫细胞中DNA疫苗的结合蛋白进行初步鉴定。 提取小鼠脾脏免疫细胞的细胞质蛋白,将细胞质蛋白分别与 DNA 疫苗 pVAX-OVA 和空载体 pVAX 共孵育,孵育后首先由琼脂糖凝胶电泳分离与DNA结合的蛋白,然后通过SDS-PAGE进行分离和纯化,最后应用质谱技术分析其蛋白组分。质谱结果初步鉴定了免疫细胞中pVAX-OVA 结合的蛋白有IQ motif containing F4 等。免疫细胞中与 pVAX 结合的蛋白有Foxl2,SUV420H2 和 gamma actin 等。Foxl2具有核定位序列,DNA结合区域和与核转运蛋白相互作用的特点,可能对DNA疫苗的进入具有促进作用。这些蛋白质是否可以影响DNA疫苗有效进入细胞核进行基因表达需要进一步研究。

Examination DNA Vaccine-binding Proteins in the Immune Cells

Examination DNA vaccine-binding proteins in the immune cells. Cytoplasmic proteins were extracted from splenocytes of mice and were incubated with DNA vaccine, pVAX-OVA as the experimental group, and empty vector pVAX, as the control group. The bound proteins were isolated by an agarose gel and SDS-PAGE and examined by MALDI-TOF mass spectrometry to analyze their sequences. In splenocytes, the potential proteins interacted with pVAX are Foxl2,SUV420H2 and gamma actin, and the potential proteins interacted with pVAX-OVA are IQ motif containing F4. Foxl2 may improve nuclear import of DNA vaccine, which includes nuclear localization signals (NLS), conserved DNA-binding forkhead domain and interaction with transporter proteins. The interactions between DNA and those proteins may or may not be beneficial for DNA vaccine entering nuclear before its expression.