A cradle for new proteins: trigger factor at the ribosome

Maier, Timm; Ferbitz, Lars; Deuerling, Elke; Ban, Nenad

doi:10.1016/j.sbi.2005.03.005

Cited by 86 publications

(71 citation statements)

References 52 publications

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“…In contrast, the ability of E. coli to correctly fold recombinant proteins containing multiple disulfide bonds is limited, due the reducing environment of the cytosol and the limited capacity for co-translational folding. 10,[30][31][32][33][34] We used combinatorial optimization methods to rapidly and predictably optimize expression of a Fab that binds to the hIL-13 α1 receptor. 25 We optimized redox conditions with added glutathione buffers and disulfide isomerase chaperone (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The covalent assembled Fab and IgG concentrations was determined from the fraction of fully assembled antibody pixels under non-reducing conditions, relative to fully reduced HC, LC and clipped product bands that correspond to the soluble protein produced, as analyzed by autoradiography using a folding. 30,32,33 Indeed, previously reported yields of Fabs are less than 50 μg/mL, 37,38 or for an IgG ca. 1 μg/mL, 6 in E. coli based cell-free systems.…”

Section: Discussionmentioning

confidence: 96%

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Aglycosylated antibodies and antibody fragments produced in a scalable in vitro transcription-translation system

Yin¹,

Garces²,

Yang³

et al. 2012

mAbs

129

157

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 96%

Aglycosylated antibodies and antibody fragments produced in a scalable in vitro transcription-translation system

Yin¹,

Garces²,

Yang³

et al. 2012

mAbs

129

157

View full text Add to dashboard Cite

show abstract

“…The C-terminal domain of TF consists of a crevice surrounded by two arm-like structures (18) similar to the N-terminal domain of the E. coli periplasmic chaperone SurA and Mycoplasma pneumoniae MPN555 (21)(22)(23). Based on photocross-linking, both arms were found to be adjacent to the nascent chain during translation (17,24).…”

mentioning

confidence: 99%

Versatility of Trigger Factor Interactions with Ribosome-Nascent Chain Complexes

Lakshmipathy

Gupta²,

Pinkert

et al. 2010

Journal of Biological Chemistry

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Trigger factor (TF) is the first molecular chaperone that interacts with nascent chains emerging from bacterial ribosomes. TF is a modular protein, consisting of an N-terminal ribosome binding domain, a PPIase domain, and a C-terminal domain, all of which participate in polypeptide binding. To directly monitor the interactions of TF with nascent polypeptide chains, TF variants were site-specifically labeled with an environmentally sensitive NBD fluorophore. We found a marked increase in TF-NBD fluorescence during translation of firefly luciferase (Luc) chains, which expose substantial regions of hydrophobicity, but not with nascent chains lacking extensive hydrophobic segments. TF remained associated with Luc nascent chains for 111 ؎ 7 s, much longer than it remained bound to the ribosomes (t1 ⁄ 2 ϳ 10 -14 s). Thus, multiple TF molecules can bind per nascent chain during translation. The Escherichia coli cytosolic proteome was classified into predicted weak and strong interactors for TF, based on the occurrence of continuous hydrophobic segments in the primary sequence. The residence time of TF on the nascent chain generally correlated with the presence of hydrophobic regions and the capacity of nascent chains to bury hydrophobicity. Interestingly, TF bound the signal sequence of a secretory protein, pOmpA, but not the hydrophobic signal anchor sequence of the inner membrane protein FtsQ. On the other hand, proteins lacking linear hydrophobic segments also recruited TF, suggesting that TF can recognize hydrophobic surface features discontinuous in sequence. Moreover, TF retained significant affinity for the folded domain of the positively charged, ribosomal protein S7, indicative of an alternative mode of TF action. Thus, unlike other chaperones, TF appears to employ multiple mechanisms to interact with a wide range of substrate proteins.

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“…TF, the first chaperone to interact with the emerging nascent chain, is a 48-kDa modular protein composed of three domains: the N-terminal domain, which mediates association with the ribosome; the peptidylprolyl-cis͞trans-isomerase (PPIase) domain; and the C-terminal domain (7). TF was shown to cooperate with the DnaK system, and the combined depletion of TF and DnaK (⌬tig⌬dnaK mutation) causes Escherichia coli cell death at Ͼ30°C as well as a massive aggregation of newly synthesized polypeptides (8).…”

mentioning

confidence: 99%

Structure of trigger factor binding domain in biologically homologous complex with eubacterial ribosome reveals its chaperone action

Baram

Pyetan²,

Sittner³

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

106

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Trigger factor (TF), the first chaperone in eubacteria to encounter the emerging nascent chain, binds to the large ribosomal subunit in the vicinity of the protein exit tunnel opening and forms a sheltered folding space. Here, we present the 3.5-Å crystal structure of the physiological complex of the large ribosomal subunit from the eubacterium Deinococcus radiodurans with the N-terminal domain of TF (TFa) from the same organism. For anchoring, TFa exploits a small ribosomal surface area in the vicinity of proteins L23 and L29, by using its ''signature motif'' as well as additional structural elements. The molecular details of TFa interactions reveal that L23 is essential for the association of TF with the ribosome and may serve as a channel of communication with the nascent chain progressing in the tunnel. L29 appears to induce a conformational change in TFa, which results in the exposure of TFa hydrophobic patches to the opening of the ribosomal exit tunnel, thus increasing its affinity for hydrophobic segments of the emerging nascent polypeptide. This observation implies that, in addition to creating a protected folding space for the emerging nascent chain, TF association with the ribosome prevents aggregation by providing a competing hydrophobic environment and may be critical for attaining the functional conformation necessary for chaperone activity.protein folding ͉ nascent chain ͉ ribosomal exit tunnel ͉ ribosomal crystallography ͉ Deinococcus radiodurans

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A cradle for new proteins: trigger factor at the ribosome

Cited by 86 publications

References 52 publications

Aglycosylated antibodies and antibody fragments produced in a scalable in vitro transcription-translation system

Aglycosylated antibodies and antibody fragments produced in a scalable in vitro transcription-translation system

Versatility of Trigger Factor Interactions with Ribosome-Nascent Chain Complexes

Structure of trigger factor binding domain in biologically homologous complex with eubacterial ribosome reveals its chaperone action

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