title: Expression, purification and evaluation of recombinant lipoprotein of Salmonella typhi as a vaccine candidate.
abstract: Lipoprotein has been reported as a vaccine candidate against many pathogenic bacteria, it plays direct role as a virulence-associated function. Here the approach is toward the expression of recombinant lipoprotein of Salmonella typhi in prokaryotic host and its evaluation as a vaccine candidate. Lipoprotein gene (lp1) was cloned in pET32a expression vector in addition to Bam HI and Hind III restriction sites, and BL21(pLysS) was used as prokaryotic expression host for transformation. Lipoprotein induction was performed by IPTG and 55 kDa (31 kDa of Gene +24 kDa of vector additional protein with His-tag) was analyzed by 12% SDS-PAGE. The recombinant lipoprotein was purified by Ni-NTA affinity chromatography due to the addition of 6X His-tag in recombinant lipoprotein. Western blot analysis using anti-His tag polyclonal antibodies confirmed the specificity of recombinant lipoprotein. Immunogenicity and protection study of recombinant lipoprotein against S. Typhi was performed in BALB/c mice. Adjuvants IFA and alum salts were used to enhance the immune response. ELISA results proved that biologically active truncated recombinant lipoprotein (31 kDa) is a suitable immunogen. Alum salts used as adjuvant was effective for long-lasting immunity.
authors.affiliation: School of Biochemical Engineering, Indian Institute of Technology (BHU), Varanasi, India. Electronic address: Anand411@gmail.com.
authors.forename: Anand
authors.initials: A
authors.lastname: Kumar
authors.affiliation: School of Biochemical Engineering, Indian Institute of Technology (BHU), Varanasi, India. Electronic address: skundu.bce@itbhu.ac.in.
authors.forename: Subir
authors.initials: S
authors.lastname: Kundu
authors.affiliation: School of Biochemical Engineering, Indian Institute of Technology (BHU), Varanasi, India. Electronic address: m.debnath.bce@itbhu.ac.in.
authors.forename: Mira
authors.initials: M
authors.lastname: Debnath Das
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression / Q000276:immunology; D054884:Host-Pathogen Interactions / Q000276:immunology; D007114:Immunization; D008074:Lipoproteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D008807:Mice, Inbred BALB C; D011994:Recombinant Proteins / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D012485:Salmonella typhi / Q000235:genetics / Q000276:immunology* / Q000502:physiology; D014435:Typhoid Fever / Q000276:immunology* / Q000382:microbiology / Q000517:prevention & control; D014436:Typhoid-Paratyphoid Vaccines / Q000276:immunology*
chemical_list: D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D008074:Lipoproteins; D011994:Recombinant Proteins; D014436:Typhoid-Paratyphoid Vaccines
keywords: Immune response; Incomplete Freund's adjuvant; Lipoprotein; Salmonella typhi; Vaccine