Ribosomal protein bL27 protects translating ribosomes from tmRNA-SmpB

16S ribosomal RNA modification drives transcript-specific translation efficiency.

It is proposed that MraW modification of 16S rRNA enhances translation efficiency in general, and that specific transcripts have evolved structural features that fine-tune protein levels that may be prevalent in bacteria which exhibit uncoupled transcription and translation.

Histidine 62 in ArfB is required for stop codon-independent peptidyl-tRNA hydrolysis on the stalled ribosome.

Translating ribosomes can stall on mRNA for various reasons, including nuclease cleavage, arrest peptide sequences or ribosome collisions. In Escherichia coli, several ribosome rescue factors act to release ribosomes stalled at the 3' end of mRNA. Among these factors, ArfB can rescue stalled ribosomes without the help of other factors. ArfB consists of an N-terminal domain containing the catalytic GGQ motif, which mediates peptidyl-tRNA hydrolysis, and a C-terminal extension that functions as a sensor for recognizing stalled ribosomes. However, how these two regions coordinate to resolve ribosome stalling remains unclear. Here, using a reconstituted translation system, we found the functional importance of histidine residues in the N-terminal domain of ArfB. In particular, histidine at position 62 in E. coli ArfB was required for stop codon-independent fMet-tRNA hydrolysis, whereas its substitution did not affect affinity for the ribosome. Furthermore, directed hydroxyl radical probing revealed that H62A mutation did not significantly alter the overall positioning of either the N-terminal domain or the C-terminal extension of ArfB on the ribosome. These findings suggest that H62 contributes to ArfB function during a step following initial ribosome binding, thereby facilitating peptidyl-tRNA hydrolysis.

RPL41 stabilizes ribosome dynamics and supports long-protein homeostasis in mammals

Abstract Ribosomal protein L41 (RPL41 or eL41) is the smallest ribosomal protein and forms the eukaryote-specific bridge, eB14, near the decoding center; however, its role in mammalian translation remains unclear. In this study, we established RPL41-deficient models of human HEK293T cells and mice to define its function. Cryo-electron microscopy revealed that RPL41 constrains intersubunit conformational dynamics without inducing major local static rearrangements. Loss of RPL41 altered A-site dynamics, slowed elongation, modestly increased amino acid misincorporation, and modestly enhanced readthrough of collision-inducing reporter sequences. Quantitative proteomic analysis suggested that these translational defects compromise long-protein homeostasis, as evidenced by increased insolubility and reduced abundance of long proteins. In vivo, Rpl41−/− mice were viable but exhibited growth retardation and decreased abundance of long proteins in tissues. Our findings reveal a conserved role for RPL41 in maintaining ribosome dynamics and translational fidelity, indicating that RPL41 supports ribosome function and long-protein homeostasis in mammals.

Rescue of ribosomal protein bL27 in Streptococcus pneumoniae TIGR4 by an alternate protease

Cryo-EM reveals that Escherichia coli tRNA-transglycosylase can bind and act upon two tRNAs

Significance tRNA base-modification is an important mechanism for the regulation of protein expression on the RNA level. Several disease-causing bacteria use the enzyme tRNA-guanine transglycosylase (TGT) to upregulate translation of virulence factors which are essential for facilitating host infection. As such, TGT is a potential drug target for diseases like shigellosis. In this work, we present the cryo-EM structure of Escherichia coli TGT, identical to the Shigella enzyme, and investigate the structural basis of its interaction with tRNA. Our work reveals that, unlike all other known TGTs, the E. coli TGT homodimer forms covalent intermediates with two tRNAs. These findings advance our understanding of TGT enzymology and provide a structural foundation for the development of antibiotics and next-generation RNA-labeling tools.

Translation factors that keep the ribosome Pro-active

SUMMARY Ribosomes produce the staggering array of proteins that perform the structural and enzymatic feats of the cell. Therefore, most of the cell’s energy goes toward producing ribosomes and the work performed by them. The work of the ribosome is relatively simple—decode mRNA codons and catalyze the formation of peptide bonds. By marching iteratively along the length of an open reading frame, a complete peptide is produced. Ribosomes catalyze the formation of peptide bonds at a rate of approximately 15 amino acids per second. However, bonds between amino acids do not form with equal efficiency, and ribosomes can become stalled at difficult-to-translate sequences. Proline is unique among the amino acids in that its side chain is covalently bonded to the peptide backbone to form a rigid ring. The rigidity of proline, and especially tracts of proline, makes it a difficult substrate for peptide bond formation, but it is also an essential motif in many protein structures. Elongation factor P (EF-P) is the star player for facilitating translation of polyproline tracts. However, recently identified factors play an important supporting role, and loss of these factors incurs a severe fitness defect in the absence of EF-P. These factors include an EF-P paralog, EfpL, as well as the ABCF ATPase YfmR/Uup and YebC2. The abundance and partial redundancy of factors that prevent ribosome stalling at polyprolines highlights the structural importance of polyproline tracts and the need to facilitate their translation. Here, we review recently identified translation factors that prevent ribosome stalling at polyprolines in bacteria.