Biomedical subjects

Marat Yusupov

Publications and source records attributed to Marat Yusupov.

4 recordsLinked to original sources

PubMed2006

Structural basis for messenger RNA movement on the ribosome.

Translation initiation is a major determinant of the overall expression level of a gene. The translation of functionally active protein requires the messenger RNA to be positioned on the ribosome such that the start/initiation codon will be read first and in the correct frame. Little is known about the molecular basis for the interaction of mRNA with the ribosome at different states of translation. Recent crystal structures of the ribosomal subunits, the empty 70S ribosome and the 70S ribosome containing functional ligands have provided information about the general organization of the ribosome and its functional centres. Here we compare the X-ray structures of eight ribosome complexes modelling the translation initiation, post-initiation and elongation states. In the initiation and post-initiation complexes, the presence of the Shine-Dalgarno (SD) duplex causes strong anchoring of the 5'-end of mRNA onto the platform of the 30S subunit, with numerous interactions between mRNA and the ribosome. Conversely, the 5' end of the 'elongator' mRNA lacking SD interactions is flexible, suggesting a different exit path for mRNA during elongation. After the initiation of translation, but while an SD interaction is still present, mRNA moves in the 3'-->5' direction with simultaneous clockwise rotation and lengthening of the SD duplex, bringing it into contact with ribosomal protein S2.

Bacterial Proteins↗

PubMed2005

Conformational transition of initiation factor 2 from the GTP- to GDP-bound state visualized on the ribosome.

Initiation of protein synthesis is a universally conserved event that requires initiation factors IF1, IF2 and IF3 in prokaryotes. IF2 is a GTPase essential for binding initiator transfer RNA to the 30S ribosomal subunit and recruiting the 50S subunit into the 70S initiation complex. We present two cryo-EM structures of the assembled 70S initiation complex comprising mRNA, fMet-tRNA(fMet) and IF2 with either a non-hydrolyzable GTP analog or GDP. Transition from the GTP-bound to the GDP-bound state involves substantial conformational changes of IF2 and of the entire ribosome. In the GTP analog-bound state, IF2 interacts mostly with the 30S subunit and extends to the initiator tRNA in the peptidyl (P) site, whereas in the GDP-bound state IF2 steps back and adopts a 'ready-to-leave' conformation. Our data also provide insights into the molecular mechanism guiding release of IF1 and IF3.

Cryoelectron Microscopy↗

PubMed2005

Bulk-solvent correction in large macromolecular structures.

The estimation of the bulk-solvent contribution to the diffraction of a macromolecular crystal makes use of a solvent mask which delimits the bulk-solvent regions in the crystal. It is shown that the way this mask is usually defined in CNS contains a bias which can lead to absurd results in the case of very large structures, where the calculations can only be made on relatively coarse grids. A modified procedure is described and applied to 70S ribosome data at 5.5 A resolution. The B factor affecting the bulk solvent is also discussed. Even in this case of very high and widely variable atomic B factors, it seems sufficient to consider a constant and isotropic B factor for the bulk solvent. This is initially set to the average value of the atomic B factor, but can be refined.

Macromolecular Substances↗

PubMed2005

Translational operator of mRNA on the ribosome: how repressor proteins exclude ribosome binding.

The ribosome of Thermus thermophilus was cocrystallized with initiator transfer RNA (tRNA) and a structured messenger RNA (mRNA) carrying a translational operator. The path of the mRNA was defined at 5.5 angstroms resolution by comparing it with either the crystal structure of the same ribosomal complex lacking mRNA or with an unstructured mRNA. A precise ribosomal environment positions the operator stem-loop structure perpendicular to the surface of the ribosome on the platform of the 30S subunit. The binding of the operator and of the initiator tRNA occurs on the ribosome with an unoccupied tRNA exit site, which is expected for an initiation complex. The positioning of the regulatory domain of the operator relative to the ribosome elucidates the molecular mechanism by which the bound repressor switches off translation. Our data suggest a general way in which mRNA control elements must be placed on the ribosome to perform their regulatory task.

Bacterial Proteins↗