Pozzi G, Oggioni M R, Manganelli R, Medaglini D, Fischetti V A, Fenoglio D, Valle M T, Kunkl A, Manca F
Department of Molecular Biology, University of Siena, Italy.
Vaccine. 1994 Sep;12(12):1071-7. doi: 10.1016/0264-410x(94)90175-9.
Our genetic system for expression of heterologous proteins on the surface of the Gram-positive bacterium Streptococcus gordonii was used to express a human T-helper epitope of HIV-1 envelope glycoprotein gp120. In previous work on the naive repertoire of human T-helper cells, it was shown that a 15-amino acid synthetic peptide of the HIV-1 gp120 sequence contained an immunodominant T-helper epitope. Synthetic DNA coding for this peptide was cloned in frame within the gene for the streptococcal surface protein M6, and the gene fusion was integrated by transformation into the chromosome of S. gordonii. The expected M6-gp120 fusion protein was found to be expressed on the surface of the recombinant streptococci. To test whether the T epitope could be recognized by T cells when expressed on the bacterial surface within the context of M6, recombinant bacteria were used as antigen in proliferation assays to stimulate the 15-amino acid-specific human T-helper clone, in the presence of autologous antigen-presenting cells. Bacteria expressing the T epitope were efficiently recognized by the T cells in culture. In proliferation assays, 10(6)-10(7) bacteria induced responses comparable to those obtained by standard amounts of synthetic peptide (0.02-0.2 micrograms). Recombinant S. gordonii, a candidate for a live vaccine vector, appeared suitable for delivering T epitopes to the immune system.
我们利用在革兰氏阳性菌戈登链球菌表面表达异源蛋白的遗传系统，来表达HIV-1包膜糖蛋白gp120的人T辅助细胞表位。在先前关于人T辅助细胞天然库的研究中发现，HIV-1 gp120序列的一个15个氨基酸的合成肽含有一个免疫显性T辅助细胞表位。编码该肽的合成DNA与链球菌表面蛋白M6的基因读框内克隆，通过转化将基因融合体整合到戈登链球菌的染色体中。发现预期的M6-gp120融合蛋白在重组链球菌表面表达。为了测试在M6背景下在细菌表面表达时T表位是否能被T细胞识别，在自体抗原呈递细胞存在的情况下，将重组细菌用作增殖试验中的抗原，以刺激15个氨基酸特异性的人T辅助细胞克隆。表达T表位的细菌在培养中能被T细胞有效识别。在增殖试验中，10⁶-10⁷个细菌诱导的反应与标准量的合成肽（0.02-0.2微克）所获得的反应相当。重组戈登链球菌作为一种活疫苗载体的候选物，似乎适合将T表位递送至免疫系统。
