Atomic insights reveal fidelity mechanisms of eukaryotic protein synthesis.

Protein synthesis involves a critical step where messenger RNA (mRNA) and transfer RNAs (tRNAs) must move in tandem to advance the mRNA reading frame by one codon. This process, known as translocation, is catalyzed by elongation factor G (EF-G) in prokaryotes and elongation factor 2 (eEF2) in archaea and eukaryotes. While eEF2 not only accelerates translocation but also maintains reading frame fidelity, high-resolution structural insights into eukaryotic translocation have remained limited compared to the extensively studied prokaryotic system.

Molecular mechanism of substrate transport by human peroxisomal ABCD3.

Peroxisomes are eukaryotic oxidative organelles involved in numerous metabolic functions that include fatty acid oxidation, bile acid synthesis, and detoxification of reactive oxygen species. ATP-binding cassette transporters of the D subfamily (ABCD1-3) mediate the import of CoA thioesters of fatty acids into the peroxisome. ABCD3, the most abundant of these transporters in the peroxisomal membrane, facilitates the transport of a broad spectrum of substrates including branched-chain fatty acids, very long-chain fatty acids, bile salt intermediates, and dicarboxylic acids.

Structural basis of BAK sequestration by MCL-1 in apoptosis.

Apoptosis controls cell fate, ensuring tissue homeostasis and promoting disease when dysregulated. The rate-limiting step in apoptosis is mitochondrial poration by the effector B cell lymphoma 2 (BCL-2) family proteins BAK and BAX, which are activated by initiator BCL-2 homology 3 (BH3)-only proteins (e.g.

The role of ATP-binding Cassette subfamily B member 6 in the inner ear.

ABCB6 has been implicated in dyschromatosis universalis hereditaria, a condition characterized by hyperpigmented and hypopigmented skin macules. Dyschromatosis universalis hereditaria can also present with hearing loss. Dyschromatosis universalis hereditaria-associated mutations in ABCB6 have been reported, but the role of this protein in the inner ear has not been studied.

Rumicidins are a family of mammalian host-defense peptides plugging the 70S ribosome exit tunnel.

The antimicrobial resistance crisis along with challenges of antimicrobial discovery revealed the vital necessity to develop new antibiotics. Many of the animal proline-rich antimicrobial peptides (PrAMPs) inhibit the process of bacterial translation. Genome projects allowed to identify immune-related genes encoding animal host defense peptides.

Low-dose cryo-electron ptychography of proteins at sub-nanometer resolution.

Cryo-transmission electron microscopy (cryo-EM) of frozen hydrated specimens is an efficient method for the structural analysis of purified biological molecules. However, cryo-EM and cryo-electron tomography are limited by the low signal-to-noise ratio (SNR) of recorded images, making detection of smaller particles challenging. For dose-resilient samples often studied in the physical sciences, electron ptychography - a coherent diffractive imaging technique using 4D scanning transmission electron microscopy (4D-STEM) - has recently demonstrated excellent SNR and resolution down to tens of picometers for thin specimens imaged at room temperature.

The archaeal highly thermostable GH35 family β-galactosidase DaβGal has a unique seven domain protein fold.

The most extensively studied β-d-galactosidases (EC3.2.1.

Cryo-EM structure of the inactive ribosome complex accumulated in chick embryo cells in cold-stress conditions.

Here, we present the high-resolution structure of the Gallus gallus 80S ribosome obtained from cold-treated chicken embryos. The translationally inactive ribosome complex contains elongation factor eEF2 with GDP, SERPINE1 mRNA binding protein 1 (SERBP1) and deacylated tRNA in the P/E position, showing common features with complexes already described in mammals. Modeling of most expansion segments of G.

mRNA reading frame maintenance during eukaryotic ribosome translocation.

One of the most critical steps of protein synthesis is coupled translocation of messenger RNA (mRNA) and transfer RNAs (tRNAs) required to advance the mRNA reading frame by one codon. In eukaryotes, translocation is accelerated and its fidelity is maintained by elongation factor 2 (eEF2). At present, only a few snapshots of eukaryotic ribosome translocation have been reported.

Structural conservation of antibiotic interaction with ribosomes.

The ribosome is a major target for clinically used antibiotics, but multidrug resistant pathogenic bacteria are making our current arsenal of antimicrobials obsolete. Here we present cryo-electron-microscopy structures of 17 distinct compounds from six different antibiotic classes bound to the bacterial ribosome at resolutions ranging from 1.6 to 2.

mRNA decoding in human is kinetically and structurally distinct from bacteria.

In all species, ribosomes synthesize proteins by faithfully decoding messenger RNA (mRNA) nucleotide sequences using aminoacyl-tRNA substrates. Current knowledge of the decoding mechanism derives principally from studies on bacterial systems. Although key features are conserved across evolution, eukaryotes achieve higher-fidelity mRNA decoding than bacteria.

Cryo-EM structures and binding of mouse and human ACE2 to SARS-CoV-2 variants of concern indicate that mutations enabling immune escape could expand host range.

Investigation of potential hosts of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is crucial to understanding future risks of spillover and spillback. SARS-CoV-2 has been reported to be transmitted from humans to various animals after requiring relatively few mutations. There is significant interest in describing how the virus interacts with mice as they are well adapted to human environments, are used widely as infection models and can be infected.

DNA-measuring Wadjet SMC ATPases restrict smaller circular plasmids by DNA cleavage.

Structural maintenance of chromosome (SMC) complexes fold DNA by loop extrusion to support chromosome segregation and genome maintenance. Wadjet systems (JetABCD/MksBEFG/EptABCD) are derivative SMC complexes with roles in bacterial immunity against selfish DNA. Here, we show that JetABCD restricts circular plasmids with an upper size limit of about 100 kb, whereas a linear plasmid evades restriction.

Two Distinct Modes of DNA Binding by an MCM Helicase Enable DNA Translocation.

A six-subunit ATPase ring forms the central hub of the replication forks in all domains of life. This ring performs a helicase function to separate the two complementary DNA strands to be replicated and drives the replication machinery along the DNA. Disruption of this helicase/ATPase ring is associated with genetic instability and diseases such as cancer.

Structural insights into the regulation of Cas7-11 by TPR-CHAT.

The CRISPR-guided caspase (Craspase) complex is an assembly of the target-specific RNA nuclease known as Cas7-11 bound to CRISPR RNA (crRNA) and an ancillary protein known as TPR-CHAT (tetratricopeptide repeats (TPR) fused with a CHAT domain). The Craspase complex holds promise as a tool for gene therapy and biomedical research, but its regulation is poorly understood. TPR-CHAT regulates Cas7-11 nuclease activity via an unknown mechanism.

Directed evolution of the rRNA methylating enzyme Cfr reveals molecular basis of antibiotic resistance.

Alteration of antibiotic binding sites through modification of ribosomal RNA (rRNA) is a common form of resistance to ribosome-targeting antibiotics. The rRNA-modifying enzyme Cfr methylates an adenosine nucleotide within the peptidyl transferase center, resulting in the C-8 methylation of A2503 (mA2503). Acquisition of results in resistance to eight classes of ribosome-targeting antibiotics.

Structure of Hsp90-Hsp70-Hop-GR reveals the Hsp90 client-loading mechanism.

Maintaining a healthy proteome is fundamental for the survival of all organisms. Integral to this are Hsp90 and Hsp70, molecular chaperones that together facilitate the folding, remodelling and maturation of the many 'client proteins' of Hsp90. The glucocorticoid receptor (GR) is a model client protein that is strictly dependent on Hsp90 and Hsp70 for activity.

Mechanism for the activation of the anaplastic lymphoma kinase receptor.

Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase (RTK) that regulates important functions in the central nervous system. The ALK gene is a hotspot for chromosomal translocation events that result in several fusion proteins that cause a variety of human malignancies. Somatic and germline gain-of-function mutations in ALK were identified in paediatric neuroblastoma.

Structural basis of early translocation events on the ribosome.

Peptide-chain elongation during protein synthesis entails sequential aminoacyl-tRNA selection and translocation reactions that proceed rapidly (2-20 per second) and with a low error rate (around 10 to 10 at each step) over thousands of cycles. The cadence and fidelity of ribosome transit through mRNA templates in discrete codon increments is a paradigm for movement in biological systems that must hold for diverse mRNA and tRNA substrates across domains of life. Here we use single-molecule fluorescence methods to guide the capture of structures of early translocation events on the bacterial ribosome.

RbfA Is Involved in Two Important Stages of 30S Subunit Assembly: Formation of the Central Pseudoknot and Docking of Helix 44 to the Decoding Center.

Ribosome biogenesis is a highly coordinated and complex process that requires numerous assembly factors that ensure prompt and flawless maturation of ribosomal subunits. Despite the increasing amount of data collected, the exact role of most assembly factors and mechanistic details of their operation remain unclear, mainly due to the shortage of high-resolution structural information. Here, using cryo-electron microscopy, we characterized 30S ribosomal particles isolated from an   strain with a deleted gene for the RbfA factor.

Structural analysis of the full-length human LRRK2.

Mutations in leucine-rich repeat kinase 2 (LRRK2) are commonly implicated in the pathogenesis of both familial and sporadic Parkinson's disease (PD). LRRK2 regulates critical cellular processes at membranous organelles and forms microtubule-based pathogenic filaments, yet the molecular basis underlying these biological roles of LRRK2 remains largely enigmatic. Here, we determined high-resolution structures of full-length human LRRK2, revealing its architecture and key interdomain scaffolding elements for rationalizing disease-causing mutations.

Delivery of functional exogenous proteins by plant-derived vesicles to human cells in vitro.

Plant-derived extracellular vesicles (EVs) gain more and more attention as promising carriers of exogenous bioactive molecules to the human cells. Derived from various edible sources, these EVs are remarkably biocompatible, biodegradable and highly abundant from plants. In this work, EVs from grapefruit juice were isolated by differential centrifugation followed by characterization of their size, quantity and morphology by nanoparticle tracking analysis, dynamic light scattering, atomic force microscopy and cryo-electron microscopy (Cryo-EM).

Structural insights into the regulation of human serine palmitoyltransferase complexes.

Sphingolipids are essential lipids in eukaryotic membranes. In humans, the first and rate-limiting step of sphingolipid synthesis is catalyzed by the serine palmitoyltransferase holocomplex, which consists of catalytic components (SPTLC1 and SPTLC2) and regulatory components (ssSPTa and ORMDL3). However, the assembly, substrate processing and regulation of the complex are unclear.

Allosteric coupling between α-rings of the 20S proteasome.

Proteasomal machinery performs essential regulated protein degradation in eukaryotes. Classic proteasomes are symmetric, with a regulatory ATPase docked at each end of the cylindrical 20S. Asymmetric complexes are also present in cells, either with a single ATPase or with an ATPase and non-ATPase at two opposite ends.