Jeffs S A, Goriup S, Stacey G, Yuen C-T, Holmes H
Division of Retrovirology, National Institute for Biological Standards and Control, Blanche Lane, South Mimms, Potters Bar, Herts, EN6 3QG, UK.
Appl Microbiol Biotechnol. 2006 Sep;72(2):279-90. doi: 10.1007/s00253-005-0256-7. Epub 2006 Jan 27.
The productivity of stable Chinese hamster ovary cell lines secreting HIV-1 monomeric (IIIB gp120) and oligomeric (UG21 gp140) recombinant envelope glycoproteins was compared in serum-containing (S+), serum-free (S-) and protein-free (P-) culture media. UG21 gp140 expression was greatest in S+ medium, while IIIBgp120 production was lower than gp140 in all three media but highest in S-. UG21 gp140 production was highest in standard 850-cm2 roller bottle cultures in S+ media, peaking after 14 days of incubation, while expression levels in the three media were 0.5 (S+), 0.4 (S-) and 0.2 (P-) mg/l, from which 90, 80 and 12% of gp140, respectively, could be purified by immunoaffinity chromatography. Purified UG21 gp140 from S+ and S- media possessed biological functionality as evidenced by CD4 and monoclonal antibody (Mab) binding. In contrast, UG21 gp140 from P- medium appears to be misfolded and non-functional. Despite the possession of a different N-linked glycan profile, UG21 gp140 from S- media shows very similar CD4 and Mab binding characteristics to S+ UG21 gp140. The relevance of these findings to HIV vaccine development is discussed.
在含血清（S+）、无血清（S-）和无蛋白（P-）的培养基中，比较了分泌HIV-1单体（IIIB gp120）和寡聚体（UG21 gp140）重组包膜糖蛋白的稳定中国仓鼠卵巢细胞系的生产力。UG21 gp140在S+培养基中的表达最高，而IIIB gp120在所有三种培养基中的产量均低于gp140，但在S-中最高。在S+培养基的标准850-cm2滚瓶培养中，UG21 gp140的产量最高，在培养14天后达到峰值，而在三种培养基中的表达水平分别为0.5（S+）、0.4（S-）和0.2（P-）mg/l，通过免疫亲和层析分别可纯化出90%、80%和12%的gp140。从S+和S-培养基中纯化的UG21 gp140具有生物学功能，CD4和单克隆抗体（Mab）结合证明了这一点。相比之下，来自P-培养基的UG21 gp140似乎错误折叠且无功能。尽管具有不同的N-连接聚糖谱，但来自S-培养基的UG21 gp140与来自S+的UG21 gp140显示出非常相似的CD4和Mab结合特性。讨论了这些发现与HIV疫苗开发的相关性。
