应用乙肝核心抗原增强肌肉抑制素免疫原性的研究

Study on the Improvement of Immunogenicity of Myostatin with Hepatitis B Core Antigen

为提高肌肉抑制素(Myostatin,MSTN)免疫原性,进行了乙肝核心抗原(HBcAg)作为分子佐剂可能性的研究。利用PCR方法获得了MSTN的C端片段,并分别在乙肝核心抗原的N端、C端及中间位点融合,构建了3个融合表达克隆:pET-30a-MSTN/HBcAg(N)、pET-30a-MSTN/HBcAg(C)和pET-30a-MSTN/HBcAg(M),转化大肠杆菌后IPTG诱导表达,然后利用Ni2+亲和层析纯化融合蛋白。应用重组蛋白免疫小鼠后利用间接ELISA法检测抗血清效价。结果表明:融合蛋白免疫效果要显著优于MSTN(P<0.05);融合蛋白MSTN/HBcAg(M)的免疫效果最佳。乙肝核心抗原可以作为分子佐剂以增强肌肉抑制素免疫原性。

Myostatin is a member of the TGF-β superfamily that functions as a negative regulator of skeletal muscle development in mammals.Targeting the myostatin pathway may be an effective strategy for increasing muscle growth.To improve immunogenicity of myostatin,the possibility of hepatitis B core antigen as molecular adjuvant was studied.First,myostatin C-domain was amplified by PCR and fused to the gene of hepatitis B core antigen at the position of N-terminal,C-terminal and internal respectively.Then three expression vector pET-30a-MSTN/HBcAg （ N）, pET-30a-MSTN/HBcAg （C） and pET-30a-MSTN/HBcAg（M）was constructed. The fused proteins were expressed in E. coli. and were purified by Ni2 ＋ affinity chromatography. Then male Kunming white mice were immunized with single fused protein or recombinant C- domain. The indirect ELISA assay was used to detect the titer of antiserum against myostatin. The results shown：The im- mune effect of fused proteins was significantly better than that of recombinant C-domain （ P 〈 0.05） ; The immune effect of fused protein MSTN/HBcAg（M）was the best in three fused protein.These results proved that HBcAg could be used as molecular adjuvant to improve the immunogenicity of myostatin.