Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:1.5.1.3 (dihydrofolate reductase)
 5,819 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The interaction of GroEL with urea-unfolded dihydrofolate reductase (DHFR) has been studied in the presence of DHFR substrates by investigating the ability of GroES to release enzyme under conditions where a stable GroES-GroEL-DHFR ternary complex can be formed. In these circumstances, GroES could only partially discharge the DHFR if ADP was present in the solution and approximately half of the DHFR remained bound on the chaperonin. This bound DHFR could be rescued by addition of ATP and KCl into the refolding mixture. The stable ternary complex did not show any significant protection of bound DHFR against proteolysis by Proteinase K. These results are in contrast to those observed with the GroEL-DHFR complex formed by thermal inactivation of DHFR at 45 degrees C in which GroES addition leads to partial protection of bound DHFR. Thus, the method of presentation influences the properties of the bound intermediates. It is suggested that the ability of GroES to bind on the same side of the GroEL double toroid as the target protein and displace it into the central cavity depends on the way the protein-substrate is presented to the GroEL molecule. Therefore, the compact folding intermediate formed by thermal unfolding can be protected against proteolysis after GroES binds to form a ternary complex. In addition, structural changes within GroEL induced by the experimental conditions may contribute to differences in the properties of the complexes. The more open urea-unfolded DHFR binds on the surface of chaperonin and can be displaced into solution by the tighter binding GroES molecule. It is suggested that the state of the unfolded protein when it is presented to GroEL determines the detailed mechanism of its assisted refolding. It follows that individual proteins, having characteristic folding intermediates, can have different detailed mechanisms of chaperonin-assisted folding.
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PMID:Conditions of forming protein complexes with GroEL can influence the mechanism of chaperonin-assisted refolding. 899 21

Although protein secretion occurs post-translationally in bacteria and is mainly a cotranslational event in Eukarya, the relationship between the translation and translocation of secreted proteins in Archaea is not known. To address this question, the signal peptide-encoding region of the surface layer glycoprotein gene from the Haloarchaea Haloferax volcanii was fused either to the cellulose-binding domain of the Clostridium thermocellum cellulosome or to the cytoplasmic enzyme dihydrofolate reductase from H. volcanii. Signal peptide-cleaved mature versions of both the cellulose-binding domain and dihydrofolate reductase could be detected in the growth medium of transformed H. volcanii cells. Immunoblot analysis revealed, however, the presence of full-length signal peptide-bearing forms of both proteins inside the cytoplasm of the transformed cells. Proteinase accessibility assays confirmed that the presence of cell-associated signal peptide-bearing proteins was not due to medium contamination. Moreover, the pulse-radiolabeled signal peptide cellulose-binding domain chimera could be chased from the cytoplasm into the growth medium even following treatment with anisomycin, an antibiotic inhibitor of haloarchaeal protein translation. Thus, these results provide evidence that, in Archaea, at least some secreted proteins are first synthesized inside the cell and only then translocated across the plasma membrane into the medium.
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PMID:Post-translational secretion of fusion proteins in the halophilic archaea Haloferax volcanii. 1256 48