High-resolution Structures of Ribosomal Subunits: Initiation, Inhibition, and Conformational Variability (original) (raw)
Molecular Cell, 2003
Noller et al., 1992; Garrett and Rodriguez-Fon-Weizmann Institute seca, 1995; Samaha et al., 1995). Crystal structures of 76100 Rehovot complexes of Thermus thermophilus ribosomes (T70S) Israel with tRNA (Yusupov et al., 2001) as well as of the large 2 Max-Planck-Research Unit for Ribosomal Structure ribosomal subunits from the archaeon Haloarcula maris-Notkestrasse 85 mortui (H50S) and the mesophilic eubacterium Deino-22603 Hamburg coccus radiodurans (D50S) with various substrate pepti-Germany dyl-transferase analogs or inhibitors (Nissen et al., 2000; 3 Max-Planck-Institute for Molecular Genetics Schluenzen et al., 2001; Schmeing et al., 2002; Hansen Ihnestrasse 73 et al., 2002) show that the PTC can be described as a 4 FB Biologie, Chemie, Pharmazie pocket with a tunnel emerging from it. It is located at Frei University Berlin the bottom of a cavity containing all of the nucleotides Takustrasse 3
Structures of the Bacterial Ribosome at 3.5 Å Resolution
Science, 2005
We describe two structures of the intact bacterial ribosome from Escherichia coli determined to a resolution of 3.5 angstroms by x-ray crystallography. These structures provide a detailed view of the interface between the small and large ribosomal subunits and the conformation of the peptidyl transferase center in the context of the intact ribosome. Differences between the two ribosomes reveal a high degree of flexibility between the head and the rest of the small subunit. Swiveling of the head of the small subunit observed in the present structures, coupled to the ratchet-like motion of the two subunits observed previously, suggests a mechanism for the final movements of messenger RNA (mRNA) and transfer RNAs (tRNAs) during translocation.
Ribosomal crystallography: Peptide bond formation and its inhibition
Biopolymers, 2003
Ribosomes, the universal cellular organelles catalyzing the translation of genetic code into proteins, are protein/RNA assemblies, of a molecular weight 2.5 mega Daltons or higher. They are built of two subunits that associate for performing protein biosynthesis. The large subunit creates the peptide bond and provides the path for emerging proteins. The small has key roles in initiating the process and controlling its fidelity.
Structure and Dynamics of the Mammalian Ribosomal Pretranslocation Complex
Molecular Cell, 2011
Although the structural core of the ribosome is conserved in all kingdoms of life, eukaryotic ribosomes are significantly larger and more complex than their bacterial counterparts. The extent to which these differences influence the molecular mechanism of translation remains elusive. Multiparticle cryo-electron microscopy and single-molecule FRET investigations of the mammalian pretranslocation complex reveal spontaneous, large-scale conformational changes, including an intersubunit rotation of the ribosomal subunits. Through structurally related processes, tRNA substrates oscillate between classical and at least two distinct hybrid configurations facilitated by localized changes in their L-shaped fold. Hybrid states are favored within the mammalian complex. However, classical tRNA positions can be restored by tRNA binding to the E site or by the eukaryotic-specific antibiotic and translocation inhibitor cycloheximide. These findings reveal critical distinctions in the structural and energetic features of bacterial and mammalian ribosomes, providing a mechanistic basis for divergent translation regulation strategies and species-specific antibiotic action.
Functional aspects of ribosomal architecture: symmetry, chirality and regulation
Journal of Physical Organic Chemistry, 2004
High-resolution structures of both ribosomal subunits revealed that most stages of protein biosynthesis, including decoding of genetic information, are navigated and controlled by the elaborate ribosomal architectural-design. Remote interactions govern accurate substrate alignment within a flexible active-site pocket [peptidyl transferase center (PTC)], and spatial considerations, due mainly to a universal mobile nucleotide, U2585, ensure proper chirality by interfering with D-amino-acids incorporation. tRNA translocation involves two correlated motions: overall mRNA/tRNA (messenger and transfer RNA) shift, and a rotation of the tRNA single-stranded aminoacylated-3′ end around the bond connecting it with the tRNA helical-regions. This bond coincides with an axis passing through a sizable symmetry-related region, identified around the PTC in all large-subunit crystal structures. Propelled by a bulged universal nucleotide, A2602, positioned at the two-fold symmetry axis, and guided by a ribosomal-RNA scaffold along an exact pattern, the rotatory motion results in stereochemistry optimal for peptide-bond formation and in geometry ensuring nascent proteins entrance into their exit tunnel. Hence, confirming that ribosomes contribute positional rather than chemical catalysis, and that peptide bond formation is concurrent with A- to P-site tRNA passage. Connecting between the PTC, the decoding center, the tRNA entrance and exit points, the symmetry-related region can transfer intra-ribosomal signals between remote functional locations, guaranteeing smooth processivity of amino acids polymerization. Ribosomal proteins are involved in accurate substrate placement (L16), discrimination and signal transmission (L22) and protein biosynthesis regulation (CTC). Residing on the exit tunnel walls near its entrance, and stretching to its opening, protein-L22 can mediate ribosome response to cellular regulatory signals, since it can swing across the tunnel, causing gating and elongation arrest. Each of the protein CTC domains has a defined task. The N-terminal domain stabilizes the intersubunit-bridge confining the A-site-tRNA entrance. The middle domain protects the bridge conformation at elevated temperatures. The C-terminal domain can undergo substantial conformational rearrangements upon substrate binding, indicating CTC participation in biosynthesis-control under stressful conditions. Copyright © 2004 John Wiley & Sons, Ltd.
Current Protein and Peptide Science, 2002
Our understanding of the process of translation has progressed rapidly since the availability of highly resolved structures for the ribosome. A wealth of information has emerged in terms of both RNA and protein structure and the interplay between them. This has prompted us to revisit the astonishing "treasure trove" of functional data regarding the ribosome that has accumulated over the past decades. Here we try a systematic synopsis of these ribosomal functions in light of the cryo-electron microscopic structures (resolution >7 Å) and the atomic x-ray structures (>2.4 Å) of the ribosome.
Decision letter: Structure of the bacterial ribosome at 2 Å resolution
2020
Using cryo-electron microscopy (cryo-EM), we determined the structure of the Escherichia coli 70S ribosome with a global resolution of 2.0 Å. The maps reveal unambiguous positioning of protein and RNA residues, their detailed chemical interactions, and chemical modifications. Notable features include the first examples of isopeptide and thioamide backbone substitutions in ribosomal proteins, the former likely conserved in all domains of life. The maps also reveal extensive solvation of the small (30S) ribosomal subunit, and interactions with A-site and P-site tRNAs, mRNA, and the antibiotic paromomycin. The maps and models of the bacterial ribosome presented here now allow a deeper phylogenetic analysis of ribosomal components including structural conservation to the level of solvation. The high quality of the maps should enable future structural analyses of the chemical basis for translation and aid the development of robust tools for cryo-EM structure modeling and refinement.
RNA, 2020
It is only after recent advances in cryo-electron microscopy that it is now possible to describe at high-resolution structures of large macromolecules that do not crystalize. Purified 30S subunits interconvert between an “active” and “inactive” conformation. The active conformation was described by crystallography in the early 2000s, but the structure of the inactive form at high resolution remains unsolved. Here we used cryo-electron microscopy to obtain the structure of the inactive conformation of the 30S subunit to 3.6 Å resolution and study its motions. In the inactive conformation, an alternative base-pairing of three nucleotides causes the region of helix 44, forming the decoding center to adopt an unlatched conformation and the 3′ end of the 16S rRNA positions similarly to the mRNA during translation. Incubation of inactive 30S subunits at 42°C reverts these structural changes. The air–water interface to which ribosome subunits are exposed during sample preparation also peel...