Publikation: Trigger factor peptidyl-prolyl cis/trans isomerase activity is not essential for the folding of cytosolic proteins in Escherichia coli
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The ribosome-associated Trigger Factor (TF) cooperates with the DnaK system to assist the folding of newly synthesized polypeptides in Escherichia coli. TF unifies two functions in one to promote proper protein folding in vitro. First, as a chaperone it binds to unfolded protein substrates, thereby preventing aggregation and supporting productive folding. Second, TF catalyzes the cis/trans isomerization of peptidyl-prolyl bonds, which can be a rate-limiting step in protein folding. Here, we investigated whether the peptidyl-prolyl cis/trans isomerase (PPIase) function is essential for the folding activity of TF in vitro and in vivo by separating these two TF activities through site-directed mutagenesis of the PPIase catalytic center. Of the four different TF variants carrying point mutations in the PPIase domain, only the exchange of the conserved residue Phe- 198 to Ala (TF F198A) abolished the PPIase activity of TF toward both a tetrapeptide and the model protein substrate RNase T1 in vitro. In contrast, all other activities of TF F198A tested were comparable with wild type TF. TF F198A retained a similar binding specificity toward membrane-bound peptides, assisted the refolding of denatured D-glyceraldehyde-3-phosphate dehydrogenase in vitro, and associated with nascent polypeptides in an in vitro transcription/translation system. Importantly, expression of the TF F198A encoding gene complemented the synthetic lethality of ΔtigΔdnaK cells and prevented global protein misfolding at temperatures between 20 and 34°C in these cells. We conclude that the PPIase activity is not required for the function of TF in folding of newly synthesized proteins.
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KRAMER, Günter, Holger PATZELT, Thomas RAUCH, Thorben A. KURZ, Sonja VORDERWÜLBECKE, Bernd BUKAU, Elke DEUERLING, 2004. Trigger factor peptidyl-prolyl cis/trans isomerase activity is not essential for the folding of cytosolic proteins in Escherichia coli. In: Journal of Biological Chemistry. 2004, 279(14), pp. 14165-14170. ISSN 0021-9258. eISSN 1083-351X. Available under: doi: 10.1074/jbc.M313635200BibTex
@article{Kramer2004Trigg-8542, year={2004}, doi={10.1074/jbc.M313635200}, title={Trigger factor peptidyl-prolyl cis/trans isomerase activity is not essential for the folding of cytosolic proteins in Escherichia coli}, number={14}, volume={279}, issn={0021-9258}, journal={Journal of Biological Chemistry}, pages={14165--14170}, author={Kramer, Günter and Patzelt, Holger and Rauch, Thomas and Kurz, Thorben A. and Vorderwülbecke, Sonja and Bukau, Bernd and Deuerling, Elke} }
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