The C-terminal domain of Escherichia coli Hfq increases the stability of the hexamer.
The Hfq (Host factor 1) polypeptide is a nucleic acid binding protein involved in the synthesis of many polypeptides. Hfq particularly affects the translation and the stability of several RNAs. In an earlier study, the use of fold recognition methods allowed us to detect a relationship between Escherichia coli Hfq and the Sm topology. This topology was further validated by a series of biophysical studies and the Hfq structure was modelled on an Sm protein. Hfq forms a beta-sheet ring-shaped hexamer. As our previous study predicted a large number of alternative conformations for the C-terminal region, we have determined whether the last 19 C-terminal residues are necessary for protein function. We find that the C-terminal truncated protein is fully capable of binding a polyadenylated RNA (K(d) of 120 pm vs. 50 pm for full-length Hfq). This result shows that the functional core of E. coli Hfq resides in residues 1-70 and confirms previous genetic studies. Using equilibrium unfolding studies, however, we find that full-length Hfq is 1.8 kcal x mol(-1) more stable than its truncated variant. Electron microscopy analysis of both truncated and full-length proteins indicates a structural rearrangement between the subunits upon truncation. This conformational change is coupled to a reduction in beta-strand content, as determined by Fourier transform infra-red. On the basis of these results, we propose that the C-terminal domain could protect the interface between the subunits and stabilize the hexameric Hfq structure. The origin of this C-terminal domain is also discussed.
Arluison
Véronique
V
Institut de Biologie Physico-Chimique CNRS UPR 9073 conventionnée avec l'université Paris 7, Paris, France.
Folichon
Marc
M
Marco
Sergio
S
Derreumaux
Philippe
P
Pellegrini
Olivier
O
Seguin
Jérôme
J
Hajnsdorf
Eliane
E
Regnier
Philippe
P
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry*; D035001:Host Factor 1 Protein / Q000737:chemistry*; D007091:Image Processing, Computer-Assisted; D007700:Kinetics; D013058:Mass Spectrometry; D008854:Microscopy, Electron; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D017550:Spectroscopy, Fourier Transform Infrared; D013816:Thermodynamics; D014508:Urea / Q000494:pharmacology
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C087373:Hfq protein, E coli; D035001:Host Factor 1 Protein; D012313:RNA; D014508:Urea