Profiles

Alexey Amunts

Alexey Amunts

Forskarassistent

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Arbetar vid Institutionen för biokemi och biofysik
Telefon 08-16 10 03
E-post amunts@dbb.su.se
Besöksadress Science for Life Laboratory, Tomtebodavägen 23, Box 1031, 171 65 Solna
Rum SciLifeLab y3
Postadress Institutionen för biokemi och biofysik 106 91 Stockholm

Forskning

Cryo-EM visualisation of bioenergetic complexes.

Our research group investigates the fundamental question of how proteins are synthesized, folded and assembled into functional multicomponent membrane complexes that drive the cellular energy production.

Living cells ultimately depend on the conversion of energy derived from foodstuff and light into the chemical form of energy. This crucial bioenergetic step is performed in the membrane systems of mitochondria and chloroplasts. Each one of these organelle types has developed dedicated ribosomes that have diverged from the cytoplasmic counterparts. While mitoribosomes synthesize proteins involved in the oxidative phosphorylation, chlororibosomes produce components driving the pohotosynthetic reactions through pigment-protein units. To dissect the mechanism and dynamics of translation, membrane insertion and bioenergetics in organelles, we use cryo-EM.

Our group determined cryo-EM structures of the human mitoribosome with mRNA, tRNAs and translation activators in 8 different functional states, as well as its assembly intermediates. It revealed unique mechanisms of mRNA binding, tRNA translocation and assembly regulation. We also determined structures of the chlororibosome with translation factors that revealed divarication of the exit tunnel and experimental evidence for convergent evolution of ribosomes from chloroplasts and mitochondria.

These studies showed that protein synthesis machineries in organelles have adopted intricate compositions and unique tasks, adding incredible complexity to the records. The achieved understanding of the architecture of these specialized systems provides now a framework to study even more sophisticated questions regarding the assembly and evolution mechanisms of the critical bioenergetic membranes that fuel life.

 

Group members

Alexander Mühleip, Researcher

Yuzuru Itoh, Postdoc

Kock Flygaard, Rasmus, Postdoc

Annemarie Perez Boerema, PhD Student

Vivek Singh, PhD Student

Victor Tobiasson, PhD Student

Fei Wu, PhD Student

Vasileios-Evripidis Kyriakidis, Research Coordinator

Patrick Cottilli, Research Assistant

Mara Laguna Castro, Research Assistant

 

Recent Highlights

SciLifeLab, Stockholm University and AstraZeneca use cryo-EM to advance biomedicine

 

Work Opportunities

"Hire very smart people. Leave them alone, but with a tea room to talk. Support them so they have time, & aren't chasing money." Max Perutz                           

If you are interested working in such environment, please contact  Alexey Amunts

 

Location

Our research group is at the Science for Life Laboratory (SciLifeLab). SciLifeLab is a joint enterprise of Swedish universities that aims to provide frontline technologies for the academic community and develop cutting-edge research programs. Situated on the expanding Stockholm biomedical campus, SciLifeLab offers the opportunity to work in an internationally competitive and synergistic environment. The center combines technical expertise with advanced knowledge of molecular biology and translational medicine.

Address: Science for Life Laboratory, Tomtebodavägen 23A, 17165 Solna.

Publikationer

I urval från Stockholms universitets publikationsdatabas
  • 2019. Anton S. Petrov (et al.). Molecular biology and evolution 36 (2), 207-219

    Mitochondrial ribosomes (mitoribosomes) are essential components of all mitochondria that synthesize proteins encoded by the mitochondrial genome. Unlike other ribosomes, mitoribosomes are highly variable across species. The basis for this diversity is not known. Here, we examine the composition and evolutionary history of mitoribosomes across the phylogenetic tree by combining three-dimensional structural information with a comparative analysis of the secondary structures of mitochondrial rRNAs (mt-rRNAs) and available proteomic data. We generate a map of the acquisition of structural variation and reconstruct the fundamental stages that shaped the evolution of the mitoribosomal large subunit and led to this diversity. Our analysis suggests a critical role for ablation and expansion of rapidly evolving mt-rRNA. These changes cause structural instabilities that are patched by the acquisition of pre-existing compensatory elements, thus providing opportunities for rapid evolution. This mechanism underlies the incorporation of mt-tRNA into the central protuberance of the mammalian mitoribosome, and the altered path of the polypeptide exit tunnel of the yeast mitoribosome. We propose that since the toolkits of elements utilized for structural patching differ between mitochondria of different species, it fosters the growing divergence of mitoribosomes.

  • 2018. Annemarie Perez Boerema (et al.).

    Oxygenic photosynthesis produces oxygen and builds a variety of organic compounds, changing the chemistry of the air, the sea and fuelling the food chain on our planet. The photochemical reactions underpinning this process in plants take place in the chloroplast. Chloroplasts evolved ~1.2 billion years ago from an engulfed primordial diazotrophic cyanobacterium, and chlororibosomes are responsible for synthesis of the core proteins driving photochemical reactions. Chlororibosomal activity is spatiotemporally coupled to the synthesis and incorporation of functionally essential co-factors, implying the presence of chloroplast-specific regulatory mechanisms and structural adaptation of the chlororibosome1,2. Despite recent structural information3,4,5,6, some of these aspects remained elusive. To provide new insights into the structural specialities and evolution, we report a comprehensive analysis of the 2.9–3.1 Å resolution electron cryo-microscopy structure of the spinach chlororibosome in complex with its recycling factor and hibernation-promoting factor. The model reveals a prominent channel extending from the exit tunnel to the chlororibosome exterior, structural re-arrangements that lead to increased surface area for translocon binding, and experimental evidence for parallel and convergent evolution of chloro- and mitoribosomes.

  • 2018. Shintaro Aibara (et al.). Journal of Visualized Experiments (140)

    The human mitochondria possess a dedicated set of ribosomes (mitoribosomes) that translate 13 essential protein components of the oxidative phosphorylation complexes encoded by the mitochondria! genome. Since all proteins synthesized by human mitoribosomes are integral membrane proteins, human mitoribosomes are tethered to the mitochondrial inner membrane during translation. Compared to the cytosolic ribosome the mitoribosome has a sedimentation coefficient of 55S, half the rRNA content, no 5S rRNA and 36 additional proteins. Therefore, a higher protein-to-RNA ratio and an atypical structure make the human mitoribosome substantially distinct from its cytosolic counterpart. Despite the central importance of the mitoribosome to life, no protocols were available to purify the intact complex from human cell lines. Traditionally, mitoribosomes were isolated from mitochondria-rich animal tissues that required kilograms of starting material. We reasoned that mitochondria in dividing HEK293-derived human cells grown in nutrient-rich expression medium would have an active mitochondrial translation, and, therefore, could be a suitable source of material for the structural and biochemical studies of the mitoribosome. To investigate its structure, we developed a protocol for large-scale purification of intact mitoribosomes from HEK cells. Herein, we introduce nitrogen cavitation method as a faster, less labor-intensive and more efficient alternative to traditional mechanical shear-based methods for cell lysis. This resulted in preparations of the mitoribosome that allowed for its structural determination to high resolution, revealing the composition of the intact human mitoribosome and its assembly intermediates. Here, we follow up on this work and present an optimized and more cost-effective method requiring only similar to 10(10) cultured HEK cells. The method can be employed to purify human mitoribosomal translating complexes, mutants, quality control assemblies and mitoribosomal subunits intermediates. The purification can be linearly scaled up tenfold if needed, and also applied to other types of cells.

  • 2017. Alan Brown (et al.). Nature Structural & Molecular Biology 24 (10), 866-869

    Mammalian mitochondrial ribosomes (mitoribosomes) have less rRNA content and 36 additional proteins compared with the evolutionarily related bacterial ribosome. These differences make the assembly of mitoribosomes more complex than the assembly of bacterial ribosomes, but the molecular details of mitoribosomal biogenesis remain elusive. Here, we report the structures of two late-stage assembly intermediates of the human mitoribosomal large subunit (mt-LSU) isolated from a native pool within a human cell line and solved by cryo-EM to similar to 3-angstrom resolution. Comparison of the structures reveals insights into the timing of rRNA folding and protein incorporation during the final steps of ribosomal maturation and the evolutionary adaptations that are required to preserve biogenesis after the structural diversification of mitoribosomes. Furthermore, the structures redefine the ribosome silencing factor (RsfS) family as multifunctional biogenesis factors and identify two new assembly factors (L0R8F8 and mt-ACP) not previously implicated in mitoribosomal biogenesis.

  • 2017. Nirupa Desai (et al.). Science 355 (6324), 528-531

    Mitochondria have specialized ribosomes (mitoribosomes) dedicated to the expression of the genetic information encoded by their genomes. Here, using electron cryomicroscopy, we have determined the structure of the 75-component yeast mitoribosome to an overall resolution of 3.3 angstroms. The mitoribosomal small subunit has been built de novo and includes 15S ribosomal RNA (rRNA) and 34 proteins, including 14 without homologs in the evolutionarily related bacterial ribosome. Yeast-specific rRNA and protein elements, including the acquisition of a putatively active enzyme, give the mitoribosome a distinct architecture compared to the mammalian mitoribosome. At an expanded messenger RNA channel exit, there is a binding platform for translational activators that regulate translation in yeast but not mammalian mitochondria. The structure provides insights into the evolution and species-specific specialization of mitochondrial translation.

  • 2017. Donna Matzov (et al.). Nature Communications 8

    Formation of 100S ribosome dimer is generally associated with translation suppression in bacteria. Trans-acting factors ribosome modulation factor (RMF) and hibernating promoting factor (HPF) were shown to directly mediate this process in E. coli. Gram-positive S. aureus lacks an RMF homolog and the structural basis for its 100S formation was not known. Here we report the cryo-electron microscopy structure of the native 100S ribosome from S. aureus, revealing the molecular mechanism of its formation. The structure is distinct from previously reported analogs and relies on the HPF C-terminal extension forming the binding platform for the interactions between both of the small ribosomal subunits. The 100S dimer is formed through interactions between rRNA h26, h40, and protein uS2, involving conformational changes of the head as well as surface regions that could potentially prevent RNA polymerase from docking to the ribosome.

  • 2017. Björn O. Forsberg (et al.). IUCrJ 4, 723-727

    The introduction of direct detectors and the automation of data collection in cryo-EM have led to a surge in data, creating new opportunities for advancing computational processing. In particular, on-the-fly workflows that connect data collection with three-dimensional reconstruction would be valuable for more efficient use of cryo-EM and its application as a sample-screening tool. Here, accelerated on-the-fly analysis is reported with optimized organization of the data-processing tools, image acquisition and particle alignment that make it possible to reconstruct the three-dimensional density of the 70S chlororibosome to 3.2 angstrom resolution within 24 h of tissue harvesting. It is also shown that it is possible to achieve even faster processing at comparable quality by imposing some limits to data use, as illustrated by a 3.7 angstrom resolution map that was obtained in only 80 min on a desktop computer. These on-the-fly methods can be employed as an assessment of data quality from small samples and extended to high-throughput approaches.

Visa alla publikationer av Alexey Amunts vid Stockholms universitet

Senast uppdaterad: 28 augusti 2019

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