Modular structure of the trigger factor required for high activity in protein folding

Zarnt, Toralf; Tradler, Thomas; Stoller, Gerlind; Schölz, Christian; Schmid, Franz X.; Fischer, Gunter

doi:10.1006/jmbi.1997.1206

Cited by 84 publications

(90 citation statements)

References 33 publications

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“…These additional domains are involved in the interaction with substrate proteins such as RCM-T1, which causes a greatly enhanced catalytic activity in the refolding of RCM-T1 compared with the small PPIases (53). In contrast to this, the additional domains of FKBP52 do not have any influence on the catalytic efficiency of prolyl-peptide bond isomerization in RCM-T1 refolding.…”

Section: Resultsmentioning

confidence: 99%

Localization of the Chaperone Domain of FKBP52

Pirkl¹,

Fischer²,

Modrow³

et al. 2001

Journal of Biological Chemistry

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FKBP52, a multidomain peptidyl prolyl cis/transisomerase (PPIase), is found in complex with the chaperone Hsp90 and the co-chaperone p23. It displays both PPIase and chaperone activity in vitro. To localize these two activities to specific regions of the protein, we created and analyzed a set of fragments of FKBP52. The PPIase activity toward both peptides and proteins is confined entirely to domain 1 (amino acids 1-148). The chaperone activity, however, resides in the C-terminal part of FKBP52, mainly in the region between amino acids 264 and 400 (domain 3). Interestingly, this domain also contains the tetratricopeptide repeats, which are responsible for the binding to C-terminal amino acids of Hsp90. Competition assays with a C-terminal Hsp90 peptide suggest that the non-native protein and Hsp90 are bound by different regions within this domain.

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Section: Resultsmentioning

confidence: 99%

Localization of the Chaperone Domain of FKBP52

Pirkl¹,

Fischer²,

Modrow³

et al. 2001

Journal of Biological Chemistry

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“…The E. coli trigger factor, an isomerase of the FKBP family, has long been suggested to possess additional chaperone activity (reviewed in Ref. 35), which is dependent on the presence of its additional flanking domains (36). FkpA is a member of the FKBP family since it shows 83% sequence identity to the consensus sequence of the FK506-binding domain, derived from an alignment of mammalian, yeast, and bacterial members of this family (1).…”

Section: Effect Of Fkpa On the Yield Of Soluble And Active 4d5mentioning

confidence: 99%

The Periplasmic Escherichia coli Peptidylprolyl cis,trans-Isomerase FkpA

Ramm

Plückthun

2000

Journal of Biological Chemistry

134

114

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We recently identified FkpA by selecting for the increased yield of antibody single-chain Fv (scFv) fragments in phage display, even of those not containing cis-prolines. We have now investigated the properties of FkpA in vitro. The peptidylprolyl cis-trans-isomerase activity of FkpA was found to be among the highest of any such enzyme with a protein substrate, yet FkpA is not able to enhance the proline-limited refolding rate of the disulfide-free hu4D5-8 scFv fragment, probably due to inaccessibility of Pro-L95. Nevertheless, the yield of the soluble and functional scFv fragment was dramatically increased in vitro in the presence of FkpA. Similar effects were observed for an scFv fragment devoid of cisprolines. We are thus forced to conclude that the observed folding-assisting function is independent of the isomerase activity of the protein. The beneficial effect of FkpA was found to be due to two components. First, FkpA interacts with early folding intermediates, thus preventing their aggregation. Additionally, it has the ability to reactivate inactive protein, possibly also by binding to a partially unfolded species that may exist in equilibrium with the aggregated form, which may thus be released on a productive pathway. These in vitro measurements therefore fully reflect the in vivo results from periplasmic overexpression of FkpA.FkpA is one of three soluble peptidylprolyl cis,trans-isomerases (PPIase) 1 in the periplasm of Escherichia coli, in addition to PpiA (RotA) and SurA (1). It was first described as being similar to the macrophage infectivity potentiator of Legionella species (2). It is homologous to the FK506-binding proteins and was subsequently shown to belong to the E regulon and thus to be inducible under stress conditions reviewed in Ref. 6). Although the fkpA deletion mutant has been shown to be viable (1, 2), FkpA has been proposed to have a general folding-assisting function in the periplasm (Refs. 1 and 5; reviewed in Ref. 6). This idea was based on an fkpA deletion causing up-regulation of E and thus stimulation of degP transcription and, in addition, FkpA's own induction by extracytoplasmic stress (5) and its ability to restore near-normal E -dependent response when overexpressed (1).In the accompanying paper (7), FkpA was identified in a selection system as a factor improving the functional expression of antibody scFv fragments in the bacterial periplasm. This was achieved by overexpressing a library of E. coli proteins in a phagemid, on which a poorly expressing antibody was encoded for phage display, and panning for functional antibody. In vivo overexpression experiments provided further evidence that the yield of functional antibody scFv fragments produced in the periplasm is increased upon coexpression of FkpA. Interestingly, the effect was also observed with antibody fragments devoid of cis-prolines (7). In antibody scFv fragments, the variable domain of the heavy chain (V H ) is connected via a linker to the variable domain of the light chain (V L ). Antibody V H domains have n...

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“…The second TF domain (aa 149-245) carries a catalytic activity as a peptidyl-prolyl cis/trans isomerase (PPIase) and has homology to the FKBP (FK506 binding protein) type of PPIases. The contributions of this domain to TF's function in de novo folding of proteins is puzzling since mutations in this domain, either point mutations diminishing the catalytic PPIase activity or deletion of the entire domain, still display chaperone activity in vivo comparable to wild type TF [71,[77][78][79]. The C-terminal domain (aa 246-432), which constitutes nearly half of the TF protein, reveals no homology on the amino acid level to any other protein [80].…”

Section: In Vivo Role Of Tf In Protein Foldingmentioning

confidence: 99%

“…The C-terminal domain (aa 246-432), which constitutes nearly half of the TF protein, reveals no homology on the amino acid level to any other protein [80]. Deletion of this domain severely decreases TF chaperone activity in vitro and in vivo [71,77,78], indicating that this domain itself participates in substrate binding. Very recently, the crystal structure of E. coli TF [81][82][83][84], as well as that of an N-terminal TF fragment bound to Haloarcula marismortui 50S, were solved [81].…”

Section: In Vivo Role Of Tf In Protein Foldingmentioning

confidence: 99%

Molecular guardians for newborn proteins: ribosome-associated chaperones and their role in protein folding

Wegrzyn

Deuerling

2005

Cell. Mol. Life Sci.

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Abstract.A central dogma in biology is the conversion of genetic information into active proteins. The biosynthesis of proteins by ribosomes and the subsequent folding of newly made proteins represent the last crucial steps in this process. To guarantee the correct folding of newly made proteins, a complex chaperone network is required in all cells. In concert with ongoing protein biosynthesis, ribosome-associated factors can interact directly with emerging nascent polypeptides to protect them from degradation or aggregation, to promote folding into their native structure, or to otherwise contribute to their folding program. Eukaryotic cells possess two major ribosome-associated systems, an Hsp70/Hsp40-based chaperone system and the functionally enigmatic NAC complex, whereas prokaryotes employ the Trigger Factor chaperone. Recent structural insights into Trigger Factor reveal an intricate cradle-like structure that, together with the exit site of the ribosome, forms a protected environment for the folding of newly synthesized proteins.

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Modular structure of the trigger factor required for high activity in protein folding

Cited by 84 publications

References 33 publications

Localization of the Chaperone Domain of FKBP52

Localization of the Chaperone Domain of FKBP52

The Periplasmic Escherichia coli Peptidylprolyl cis,trans-Isomerase FkpA

Molecular guardians for newborn proteins: ribosome-associated chaperones and their role in protein folding

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