We used electron cryotomography to study the molecular arrangement of large respiratory chain complexes in mitochondria from bovine heart, potato, and three types fungi. Long rows ATP synthase dimers were observed intact cristae membrane fragments all species that examined. The dimer found exclusively on tightly curved edges. distance between along varied, but within F(1) heads was constant. angle monomers 70° or above. Complex I appeared as L-shaped densities tomograms reconstituted...

Abstract We used cryoelectron tomography to reveal the arrangements of photosystem II (PSII) and ATP synthase in vitreous sections intact chloroplasts plunge-frozen suspensions isolated thylakoid membranes. found that stroma grana thylakoids are connected at margins by staggered lamellar membrane protrusions. The stacking repeat membranes frozen-hydrated is 15.7 nm, with a 4.5-nm lumenal space 3.2-nm distance between flat stromal surfaces. chloroplast confined minimally curved regions end...

Aging is one of the most fundamental, yet least understood biological processes that affect all forms eukaryotic life. Mitochondria are intimately involved in aging, but underlying molecular mechanisms largely unknown. Electron cryotomography whole mitochondria from aging model organism Podospora anserina revealed profound age-dependent changes membrane architecture. With increasing age, typical cristae disappear and inner vesiculates. The ATP synthase dimers form rows at tips dissociate...

Surface layers (S-layers) are resilient two-dimensional protein lattices that encapsulate many bacteria and most archaea. In archaea, S-layers usually form the only structural component of cell wall thus act as final frontier between its environment. Therefore, crucial for supporting microbial life. Notwithstanding their importance, little is known about archaeal at atomic level. Here, we combined single-particle cryo electron microscopy, tomography, Alphafold2 predictions to generate an...

The archaellum is the macromolecular machinery that Archaea use for propulsion or surface adhesion, enabling them to proliferate and invade new territories. molecular composition of motor drives it appears be entirely distinct from functionally equivalent bacterial flagellum flagellar motor. Yet, structure scarcely known. Using combined modes electron cryo-microscopy (cryoEM), we have solved Pyrococcus furiosus filament at 4.2 Å resolution visualise architecture organisation its complex in...

Abstract Phages are viruses that infect bacteria and dominate every ecosystem on our planet. As well as impacting microbial ecology, physiology evolution, phages exploited tools in molecular biology biotechnology. This is particularly true for the Ff (f1, fd or M13) phages, which represent a widely distributed group of filamentous viruses. Over nearly five decades, Ffs have seen an extraordinary range applications, yet complete structure phage capsid consequently mechanisms infection...

The majority of outer membrane proteins (OMPs) from Gram-negative bacteria and many mitochondria chloroplasts are β-barrels. Insertion assembly these catalyzed by the Omp85 protein family in a seemingly conserved process. All members this exhibit characteristic N-terminal polypeptide-transport–associated (POTRA) C-terminal 16-stranded β-barrel domain. In plants, two phylogenetically distinct essential Omp85's exist chloroplast membrane, namely Toc75-III Toc75-V. Whereas Toc75-V, similar to...

A decisive step in a virus infection cycle is the recognition of specific receptor present on host cell surface, subsequently leading to delivery viral genome into interior. Until now, early stages have not been thoroughly investigated for any infecting hyperthermophilic archaea. Here, we first study focusing primary interactions between archaeal rod-shaped Sulfolobus islandicus 2 (SIRV2) (family Rudiviridae) and its hyperthermoacidophilic host, S. islandicus. We show that SIRV2 adsorption...

Abstract Type IV pili are flexible filaments on the surface of bacteria, consisting a helical assembly pilin proteins. They involved in bacterial motility (twitching), adhesion, biofilm formation and DNA uptake (natural transformation). Here, we use cryo-electron microscopy mass spectrometry to show that bacterium Thermus thermophilus produces two forms type pilus (‘wide’ ‘narrow’), differing structure protein composition. Wide composed major PilA4, while narrow so-far uncharacterized which...

Abstract Archaea use a molecular machine, called the archaellum, to swim. The archaellum consists of an ATP-powered intracellular motor that drives rotation extracellular filament composed multiple copies proteins named archaellins. In many species, several archaellin homologs are encoded in same operon; however, previous structural studies indicated filaments mainly consist only one protein species. Here, we electron cryo-microscopy elucidate structure from Methanocaldococcus villosus at...

Abstract Amongst the major types of archaeal filaments, several have been shown to closely resemble bacterial homologues Type IV pili (T4P). Within Sulfolobales , member species encode for three T4P, namely archaellum, UV-inducible pilus system (Ups) and adhesive (Aap). Whereas archaellum functions primarily in swimming motility, Ups UV-induced cell aggregation DNA-exchange, Aap plays an important role adhesion twitching motility. Here, we present a cryoEM structure model organism Sulfolobus...

The outer-membrane of Gram-negative bacteria is critical for surface adhesion, pathogenicity, antibiotic resistance and survival. major constituent – hydrophobic β-barrel O uter- M embrane P roteins (OMPs) are first secreted across the inner-membrane through Sec-translocon delivery to periplasmic chaperones, example SurA, which prevent aggregation. OMPs then offloaded β- B arrel A ssembly achinery (BAM) in insertion folding. We show H olo- T rans L ocon (HTL) an assembly protein-channel...

Mitochondrial ATP synthases form rows of dimers, which induce membrane curvature to give cristae their characteristic lamellar or tubular morphology. The angle formed between the central stalks synthase dimers varies species. Using cryo-electron tomography and sub-tomogram averaging, we determined structure dimer from nematode worm C. elegans show that differs previously structures. consequences this species-specific difference at interface were investigated by comparing S. cerevisiae...

Significance The Sulfolobus islandicus rod-shaped virus 2 (SIRV2) has developed unique mechanisms to penetrate the plasma membrane and S-layer of its host in order leave cell after replication. SIRV2 encodes 10-kDa protein PVAP, which assembles into sevenfold symmetric virus-associated pyramids (VAPs) membrane. Toward end viral replication cycle, these VAPs open form pores through S-layer, allowing egress. Here we show that PVAP inserts spontaneously forms any kind biological By electron...

Surface protein layers (S-layers) often form the only structural component of archaeal cell wall and are therefore important for survival. S-layers have a plethora cellular functions including maintenance shape, osmotic, mechanical stability, formation semipermeable protective barrier around cell, cell–cell interaction, as well surface adhesion. Despite central importance life, their 3-dimensional (3D) architecture is still poorly understood. Here we present detailed 3D electron...

The 30 kDa N-BAR domain of the human Bin1 protein is essential for generation skeletal muscle T-tubules. By electron cryo-microscopy and cryo-tomography with a direct detector, we found that Bin1-N-BAR domains assemble into scaffolds low long-range order form flexible membrane tubules. diameter tubules closely matches curved shape domain, which depends on composition target membrane. These insights are fundamental to our understanding T-tubule formation function in muscle.

Cyclophilin D (CYPD) is a mitochondrial peptidyl prolyl-cis,trans-isomerase involved in opening of the permeability transition pore (mPTP). CYPD abundance increases during aging mammalian tissues and model organism Podospora anserina. Here, we show that treatment P. anserina wild-type with low concentrations cyclophilin inhibitor cyclosporin A (CSA) extends lifespan. Transgenic strains overexpressing PaCypD are characterized by reduced stress tolerance, suffer from pronounced dysfunction...

Electron cryo-tomography is a powerful tool in structural biology, capable of visualizing the three-dimensional structure biological samples, such as cells, organelles, membrane vesicles, or viruses at molecular detail. To achieve this, aqueous sample rapidly vitrified liquid ethane, which preserves it close-to-native, frozen-hydrated state. In electron microscope, tilt series are recorded nitrogen temperature, from 3D tomograms reconstructed. The signal-to-noise ratio tomographic volume...

The uncultivated "Candidatus Altiarchaeum hamiconexum" (formerly known as SM1 Euryarchaeon) carries highly specialized nano-grappling hooks ("hami") on its cell surface. Until now little is about the major protein forming these structured fibrous surface appendages, genes involved or membrane anchoring of filaments. These aspects were analyzed in depth this study using environmental transcriptomics combined with imaging methods. Since a laboratory culture archaeon not yet available, natural...

Abstract The Ff family of filamentous bacteriophages infect gram-negative bacteria, but do not cause lysis their host cell. Instead, new virions are extruded via the phage-encoded pIV protein, which has homology with bacterial secretins. Here, we determine structure from f1 bacteriophage at 2.7 Å resolution by cryo-electron microscopy, first near-atomic a phage secretin. Fifteen subunits assemble to form gated channel in outer membrane, associated soluble domains projecting into periplasm....

Pili are filamentous surface extensions that play roles in bacterial and archaeal cellular processes such as adhesion, biofilm formation, motility, cell-cell communication, DNA uptake horizontal gene transfer. The model archaeaon Sulfolobus acidocaldarius assembles three filaments of the type-IV pilus superfamily (archaella, adhesion pili UV-inducible pili), well a so-far uncharacterised fourth filament, named "thread". Here, we report on cryo-EM structure thread. filament is highly...

Abstract Translational control is an essential process for the cell to adapt varying physiological or environmental conditions. To survive adverse conditions such as low nutrient levels, translation can be shut down almost entirely by inhibiting ribosomal function. Here we investigated eukaryotic hibernating ribosomes from microsporidian parasite Spraguea lophii in situ a combination of electron cryo-tomography and single-particle cryo-microscopy. We show that spores contain are locked...