PMID:11257542
| Citation |
Al-Herran, S and Ashraf, W (2001) The immunological dissection of the Escherichia coli molecular chaperone DnaJ. FEMS Microbiol. Lett. 196:19-23 |
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| Abstract |
In Escherichia coli, one of the main molecular chaperones is DnaJ (hsp40), which mediates in a variety of highly conserved cellular processes including protein-folding reactions and the assembly/disassembly of protein complexes. DnaJ is characterised by the presence of four distinct domains: the J-domain, glycine/phenylalanine-rich (G/F), cysteine-rich (Zn-finger) and C-terminal regions. Truncated DnaJ polypeptides (DnaJ 1-108, DnaJ Delta1-108, DnaJ Delta1-199) representing these domains were over-produced and used as a source of immunogens for the generation of sequence-specific polyclonal antibodies. Epitope mapping was achieved by Western blotting, which demonstrated the presence of antibodies directed against these domains. These characterised affinity-purified antibodies were then used to assess the role of DnaJ in the protection of firefly luciferase from irreversible heat-inactivation. In this study we have demonstrated the involvement of J-, G/F and Zn-finger domains in the protection of luciferase from heat-inactivation. The C-terminal region had only partial involvement in luciferase protection. |
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| Keywords |
Amino Acid Motifs; Antibodies/immunology; Antibodies/isolation & purification; Blotting, Western; Chromatography, Affinity; Epitope Mapping; Escherichia coli; Escherichia coli Proteins; HSP40 Heat-Shock Proteins; Heat-Shock Proteins/antagonists & inhibitors; Heat-Shock Proteins/chemistry; Heat-Shock Proteins/immunology; Heat-Shock Proteins/metabolism; Hot Temperature; Luciferases/chemistry; Luciferases/metabolism; Molecular Chaperones/antagonists & inhibitors; Molecular Chaperones/chemistry; Molecular Chaperones/immunology; Molecular Chaperones/metabolism; Protein Denaturation; Protein Structure, Tertiary; Zinc Fingers |
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