Cloning and periplasmic expression of peptidoglycan-associated lipoprotein (PAL) protein of Legionella pneumophila in Escherichia coli
Journal Title: Jundishapur Journal of Microbiology - Year 2010, Vol 3, Issue 1
Abstract
Introduction and objective: Legionella pneumophila, the etiological agent of Legionnaires’ disease, is an important cause of both community-acquired and nosocomial pneumonia; therefore, rapid diagnosis and early antibiotic treatment of pneumonia are required. Urinary antigen testing to detect Legionella antigen has proven to be the most powerful diagnostic method. Peptidoglycan-associated lipoprotein (PAL) protein of L. pneumophila, as a component of Legionella antigens, will be detected efficiently by the PAL antigen capture assay and is considered as useful diagnostic antigen to diagnose Legionella infection. Because of the transfer of protein to the periplasmic region of Escherichia coli has numerous advantages including separation from cytoplasmic proteins and the concentration of recombinant proteins in periplasm, the aim of this study was to produce periplasmic PAL protein of L. pneumophila in E. coli. Materials and methods: The pal gene of L. pneumophila serogroup 1 was amplified with specific primers, cloned and expressed under pelB signal sequence and T7 lac promoter in pET26b+ plasmid. Results: The cloning was confirmed with digestion and sequencing of recombinant pET-26b-pal plasmid. The expression of r-PAL protein in cytoplasm and periplasmic space of E. coli was approved by SDS-PAGE and western blotting. Conclusion: The results of this study demonstrated that the r-PAL protein successfully expressed in E. coli.
Authors and Affiliations
Abolfazl Gholipour, Mojtaba Moosavian, Hamid Galehdari, Manoochehr Makvandi, Amirhoushang Alvandi
Keywords
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