title: Cloning, protein expression and display of synthetic multi-epitope mycobacterial antigens on Salmonella typhi Ty21a cell surface.
abstract: Expressing proteins of interest as fusion to proteins of bacterial envelope is a powerful technique for biotechnological and medical applications. The synthetic gene (VacII) encoding for T-cell epitopes of selected genes of Mycobacterium tuberculosis namely, ESAT6, MTP40, 38 kDa, and MPT64 was fused with N- terminus of Pseudomonas syringae ice nucleation protein (INP) outer membrane protein. The fused genes were cloned into a bacterial expression vector pKK223-3. The recombinant protein was purified by Ni-NAT column. VacII gene was displayed on the cell surface of Salmonella typhi Ty21a using N-terminal region of ice nucleation proteins (INP) as an anchoring motif. Glycine method confirmed that VacII was anchored on the cell surface. Western blot analysis further identified the synthesis of INP derivatives containing the N-terminal domain INP- VacII fusion protein of the expected size (52 kDa).
authors.affiliation: Department of Biology, College of Science, King Khalid University, 61413 Abha Saudi Arabia. mohammed_sarhan@yahoo.com
authors.forename: Mohammed A A
authors.initials: MA
authors.lastname: Sarhan
authors.forename: Mustaffa
authors.initials: M
authors.lastname: Musa
authors.forename: Zainul F
authors.initials: ZF
authors.lastname: Zainuddin
mesh_terms: D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics; D001483:Base Sequence; D001709:Biotechnology / Q000379:methods*; D015153:Blotting, Western; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D018984:Epitopes, T-Lymphocyte / Q000737:chemistry / Q000235:genetics*; D015870:Gene Expression; D005813:Genes, Synthetic; D008969:Molecular Sequence Data; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D012485:Salmonella typhi / Q000235:genetics* / Q000378:metabolism
chemical_list: D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D018984:Epitopes, T-Lymphocyte; D011993:Recombinant Fusion Proteins; C063113:ice nucleation protein; C072681:Mycobacterium tuberculosis antigens