Hertle R, Mrsny R, Fitzgerald D J
Biotherapy Section, Laboratory of Molecular Biology, CCR, National Cancer Institute, Bethesda, Maryland 20892-4255, USA.
Infect Immun. 2001 Nov;69(11):6962-9. doi: 10.1128/IAI.69.11.6962-6969.2001.
Pseudomonas aeruginosa is the major infectious agent of concern for cystic fibrosis patients. Strategies to prevent colonization by this bacterium and/or neutralize its virulence factors are clearly needed. Here we characterize a dual-function vaccine designed to generate antibodies to reduce bacterial adherence and to neutralize the cytotoxic activity of exotoxin A. To construct the vaccine, key sequences from type IV pilin were inserted into a vector encoding a nontoxic (active-site deletion) version of exotoxin A. The chimeric protein, termed PE64Delta553pil, was expressed in Escherichia coli, refolded to a near-native conformation, and then characterized by various biochemical and immunological assays. PE64Delta553pil bound specifically to asialo-GM1, and, when injected into rabbits, produced antibodies that reduced bacterial adherence and neutralized the cell-killing activity of exotoxin A. Results support further evaluation of this chimeric protein as a vaccine to prevent Pseudomonas colonization in susceptible individuals.
铜绿假单胞菌是囊性纤维化患者主要关注的感染病原体。显然需要采取策略来预防这种细菌的定植和/或中和其毒力因子。在此，我们对一种双功能疫苗进行了表征，该疫苗旨在产生抗体以减少细菌黏附并中和外毒素A的细胞毒性活性。为构建该疫苗，将IV型菌毛的关键序列插入到编码外毒素A无毒（活性位点缺失）版本的载体中。这种嵌合蛋白称为PE64Delta553pil，在大肠杆菌中表达，重折叠成接近天然的构象，然后通过各种生化和免疫测定进行表征。PE64Delta553pil特异性结合去唾液酸GM1，当注射到兔子体内时，产生的抗体可减少细菌黏附并中和外毒素A的细胞杀伤活性。结果支持进一步评估这种嵌合蛋白作为预防易感个体中铜绿假单胞菌定植的疫苗。
