The expression of eukaryotic genes is precisely controlled by specific interactions between general transcription initiation factors and gene‐specific transcriptional activators. The general transcription factor TFIID, which plays an essential role in mediating transcriptional activation, is a multisubunit complex comprising the TATA box‐binding protein (TBP) and multiple TBP‐associated factors (TAFs). On the other hand, biochemical and genetic approaches have shown that the promoter‐specific transcriptional activator Sp1 has the ability to interact with one of the components of TFIID, the TBP‐associated factor TAF4. We herein report the structural details of the glutamine‐rich domains (Q‐domains) of Sp1 and TAF4 using circular dichroism (CD) and heteronuclear magnetic resonance (NMR) spectroscopy. We found that the two Q‐domains of Sp1 and four Q‐domains of TAF4 were disordered under physiological conditions. We also quantitatively analyzed the interaction between the Q‐domains of Sp1 and TAF4 by NMR and surface plasmon resonance, and detected a weak but specific association between them. Nevertheless, a detailed analysis of CD spectra suggested that any significant conformational change did not occur concomitantly with this association, at least at the level of the overall secondary structure. These results may represent a prominent and exceptional binding mode for the IDPs, which are not categorized in a well‐accepted concept of “coupled folding and binding.”

Chaperonin GroEL from Escherichia coli consists of two heptameric rings stacked back-to-back to form a cage-like structure. It assists in the folding of substrate proteins in concert with the co-chaperonin, GroES, by incorporating them into its large cavity. The mechanism underlying the incorporation of substrate proteins currently remains unclear. The flexible C-terminal residues of GroEL, which are invisible in the X-ray crystal structure, have recently been suggested to play a key role in the efficient encapsulation of substrates. These C-terminal regions have also been suggested to separate the double rings of GroEL at the bottom of the cavity. In order to elucidate the role of the C-terminal regions of GroEL on the efficient encapsulation of substrate proteins, we herein investigated the effects of C-terminal truncation on GroE-mediated folding using the green fluorescent protein (GFP) as a substrate. We demonstrated that the yield of in-cage folding mediated by a single-ring GroEL (SR1) was markedly decreased by truncation, whereas that mediated by a double-ring football-shaped complex was not affected. These results suggest that the C-terminal region of GroEL functions as a barrier between rings, preventing the leakage of GFP through the bottom space of the cage. We also found that once GFP folded into its native conformation within the cavity of SR1, it never escaped, even in the absence of the C-terminal tails. This suggests that GFP molecules escaped through the pore only when they adopted a denatured conformation. Therefore, the folding and escape of GFP from C-terminal-truncated SR1/GroES appeared to be competing with each other.