@article{22148,
  abstract     = {How the twin-arginine translocase (Tat) system transports fully folded substrate proteins across cellular membranes without disrupting membrane integrity has been a fundamental question in cell biology for decades. The Tat system, found in prokaryotes and plant organelles, recognizes a cargo signal peptide via a conserved twin-arginine motif. The multi-subunit Tat complex facilitates the proton-motive-force-dependent translocation process, yet its overall architecture has remained unknown. Here, we present the cryo-electron microscopy (cryo-EM) structure of the Escherichia coli (E. coli) trimeric TatB₃C₃ complex with bound substrate SufI, assembled in vivo. The complex adopts an unusual, wide-open, bowl-shaped architecture with a polar inner cavity. Unexpectedly, the cargo is engaged in a dual-contact mode: while the signal peptide binds inside one TatBC unit, the folded domain docks tightly onto an adjacent unit, possibly performing a proofreading function. This structure provides a mechanistic framework for substrate engagement and suggests the direct involvement of the entire Tat complex in substrate translocation.},
  author       = {Zhao, Ziyu and Sazanov, Leonid A},
  issn         = {1097-4164},
  journal      = {Molecular Cell},
  publisher    = {Elsevier},
  title        = {{Structure of E. Coli twin-arginine translocase (Tat) complex with bound cargo}},
  doi          = {10.1016/j.molcel.2026.05.026},
  year         = {2026},
}

@article{18987,
  abstract     = {Biallelic variants in NADH (nicotinamide adenine dinucleotide (NAD) + hydrogen (H))-ubiquinone oxidoreductase 1 alpha subcomplex 13 have been linked to mitochondrial complex I deficiency, nuclear type 28, based on three affected individuals from two families. With only two families reported, the clinical and molecular spectrum of NADH-ubiquinone oxidoreductase 1 alpha subcomplex 13–related diseases remains unclear. We report 10 additional affected individuals from nine independent families, identifying four missense variants (including recurrent c.170G > A) and three ultra-rare or novel predicted loss-of-function biallelic variants. Updated clinical–radiological data from previously reported families and a literature review compiling clinical features of all reported patients with isolated complex I deficiency caused by 43 genes encoding complex I subunits and assembly factors are also provided. Our cohort (mean age 7.8 ± 5.4 years; range 2.5–18) predominantly presented a moderate-to-severe neurodevelopmental syndrome with oculomotor abnormalities (84%), spasticity/hypertonia (83%), hypotonia (69%), cerebellar ataxia (66%), movement disorders (58%) and epilepsy (46%). Neuroimaging revealed bilateral symmetric T2 hyperintense substantia nigra lesions (91.6%) and optic nerve atrophy (66.6%). Protein modeling suggests missense variants destabilize a critical junction between the hydrophilic and membrane arms of complex I. Fibroblasts from two patients showed reduced complex I activity and compensatory complex IV activity increase. This study characterizes NADH-ubiquinone oxidoreductase 1 alpha subcomplex 13–related disease in 13 individuals, highlighting genotype–phenotype correlations.},
  author       = {Kaiyrzhanov, Rauan and Thompson, Kyle and Efthymiou, Stephanie and Mukushev, Askhat and Zharylkassyn, Akbota and Prasad, Chitra and Karimiani, Ehsan Ghayoor and Alvi, Javeria Raza and Niyazov, Dmitriy and Alahmad, Ahmad and Babaei, Meisam and Tajsharghi, Homa and Albash, Buthaina and Alaqeel, Ahmad and Charif, Majida and Hashemi, Narges and Heidari, Morteza and Kalantar, Seyed Mehdi and Lenaers, Guy and Mehrjardi, Mohammad Yahya Vahidi and Srinivasan, Varunvenkat M. and Gowda, Vykuntaraju K. and Mirabutalebi, Seyed Hamidreza and Carere, Deanna Alexis and Movahedinia, Mojtaba and Murphy, David and Mcfarland, Robert and Abdel-Hamid, Mohamed S. and Elhossini, Rasha M. and Alavi, Shahryar and Napier, Melanie and Belanger-Quintana, Amaya and Prasad, Asuri N. and Jakobczyk, Jessica and Roubertie, Agathe and Rupar, Tony and Sultan, Tipu and Toosi, Mehran Beiraghi and Sazanov, Leonid A and Severino, Mariasavina and Houlden, Henry and Taylor, Robert W. and Maroofian, Reza},
  issn         = {2632-1297},
  journal      = {Brain Communications},
  number       = {1},
  publisher    = {Oxford University Press},
  title        = {{Biallelic NDUFA13 variants lead to a neurodevelopmental phenotype with gradual neurological impairment}},
  doi          = {10.1093/braincomms/fcae453},
  volume       = {7},
  year         = {2025},
}

@article{19529,
  abstract     = {NRF2 is a transcription factor responsible for coordinating the expression of over a thousand cytoprotective genes. Although NRF2 is constitutively expressed, its stability is modulated by the redox-sensitive protein KEAP1 and other conditional binding partner regulators. The new era of NRF2 research has highlighted the cooperation between NRF2 and PIN1 in modifying its cytoprotective effect. Despite numerous studies, the understanding of the PIN1-NRF2 interaction remains limited. Herein, we described the binding interaction of PIN1 and three different 14-mer long phospho-peptides mimicking NRF2 protein using computer-based, biophysical, and biochemical approaches. According to our computational analyses, the residues positioned in the WW domain of PIN1 (Ser16, Arg17, Ser18, Tyr23, Ser32, Gln33, and Trp34) were found to be crucial for PIN1-NRF2 interactions. Biophysical FP assays were used to verify the computational prediction. The data demonstrated that Pintide, a peptide predominantly interacting with the PIN1 WW-domain, led to a significant reduction in the binding affinity of the NRF2 mimicking peptides. Moreover, we evaluated the impact of known PIN1 inhibitors (juglone, KPT-6566, and EGCG) on the PIN1-NRF2 interaction. Among the inhibitors, KPT-6566 showed the most potent inhibitory effect on PIN1-NRF2 interaction within an IC<jats:sub>50</jats:sub> range of 0.3–1.4 µM. Furthermore, our mass spectrometry analyses showed that KPT-6566 appeared to covalently modify PIN1 via conjugate addition, rather than disulfide exchange of the sulfonyl-acetate moiety. Altogether, such inhibitors would also be highly valuable molecular probes for further investigation of PIN1 regulation of NRF2 in the cellular context and potentially pave the way for drug molecules that specifically inhibit the cytoprotective effects of NRF2 in cancer.},
  author       = {Ozleyen, Adem and Duran, Gizem Nur and Dönmez, Serhat and Ozbil, Mehmet and Doveston, Richard G. and Tumer, Tugba Boyunegmez},
  issn         = {2045-2322},
  journal      = {Scientific Reports},
  publisher    = {Springer Nature},
  title        = {{Identification and inhibition of PIN1-NRF2 protein–protein interactions through computational and biophysical approaches}},
  doi          = {10.1038/s41598-025-89342-0},
  volume       = {15},
  year         = {2025},
}

@article{20002,
  abstract     = {While the most widely used CRISPR-Cas enzyme is the Cas9 endonuclease from Streptococcus pyogenes (Cas9), it exhibits single-turnover enzyme kinetics which leads to long residence times on product DNA. This blocks access to DNA repair machinery and acts as a major bottleneck during CRISPR-Cas9 gene editing. Cas9 can eventually be removed from the product by extrinsic factors, such as translocating polymerases, but the mechanisms contributing to Cas9 dissociation following cleavage remain poorly understood. Here, we employ truncated guide RNAs as a strategy to weaken PAM-distal nucleic acid interactions and promote faster enzyme turnover. Using kinetics-guided cryo-EM, we examine the conformational landscape of a multi-turnover Cas9, including the first detailed snapshots of Cas9 dissociating from product DNA. We discovered that while the PAM-distal product dissociates from Cas9 following cleavage, tight binding of the PAM-proximal product directly inhibits re-binding of new targets. Our work provides direct evidence as to why Cas9 acts as a single-turnover enzyme and will guide future Cas9 engineering efforts.},
  author       = {Kiernan, Kaitlyn and Taylor, David W.},
  issn         = {2041-1723},
  journal      = {Nature Communications},
  publisher    = {Springer Nature},
  title        = {{Visualization of a multi-turnover Cas9 after product release}},
  doi          = {10.1038/s41467-025-60668-7},
  volume       = {16},
  year         = {2025},
}

@article{17884,
  abstract     = {Human T cell leukemia virus type 1 (HTLV-1) immature particles differ in morphology from other retroviruses, suggesting a distinct way of assembly. Here we report the results of cryo-electron tomography studies of HTLV-1 virus-like particles assembled in vitro, as well as derived from cells. This work shows that HTLV-1 uses a distinct mechanism of Gag–Gag interactions to form the immature viral lattice. Analysis of high-resolution structural information from immature capsid (CA) tubular arrays reveals that the primary stabilizing component in HTLV-1 is the N-terminal domain of CA. Mutagenesis analysis supports this observation. This distinguishes HTLV-1 from other retroviruses, in which the stabilization is provided primarily by the C-terminal domain of CA. These results provide structural details of the quaternary arrangement of Gag for an immature deltaretrovirus and this helps explain why HTLV-1 particles are morphologically distinct.},
  author       = {Obr, Martin and Percipalle, Mathias and Chernikova, Darya and Yang, Huixin and Thader, Andreas and Pinke, Gergely and Porley, Dario J and Mansky, Louis M. and Dick, Robert A. and Schur, Florian KM},
  issn         = {1545-9985},
  journal      = {Nature Structural & Molecular Biology},
  pages        = {268--276},
  publisher    = {Springer Nature},
  title        = {{Distinct stabilization of the human T cell leukemia virus type 1 immature Gag lattice}},
  doi          = {10.1038/s41594-024-01390-8},
  volume       = {32},
  year         = {2025},
}

@article{18170,
  abstract     = {This study presents a graphene field-effect transistor (gFET) biosensor with dual detection capabilities for SARS-CoV-2: one RNA detection assay to confirm viral positivity and the other for nucleocapsid (N-)protein detection as a proxy for infectiousness of the patient. This technology can be rapidly adapted to emerging infectious diseases, making an essential tool to contain future pandemics. To detect viral RNA, the highly conserved E-gene of the virus was targeted, allowing for the determination of SARS-CoV-2 presence or absence using nasopharyngeal swab samples. For N-protein detection, specific antibodies were used. Tested on 213 clinical nasopharyngeal samples, the gFET biosensor showed good correlation with RT-PCR cycle threshold values, proving its high sensitivity in detecting SARS-CoV-2 RNA. Specificity was confirmed using 21 pre-pandemic samples positive for other respiratory viruses. The gFET biosensor had a limit of detection (LOD) for N-protein of 0.9 pM, establishing a foundation for the development of a sensitive tool for monitoring active viral infection. Results of gFET based N-protein detection corresponded to the results of virus culture in all 16 available clinical samples and thus it also proved its capability to serve as a proxy for infectivity. Overall, these findings support the potential of the gFET biosensor as a point-of-care device for rapid diagnosis of SARS-CoV-2 infection and indirect assessment of infectiousness in patients, providing additional information for clinical and public health decision-making.},
  author       = {Herdina, Anna Nele and Bozdogan, Anil and Aspermair, Patrik and Dostalek, Jakub and Klausberger, Miriam and Lingg, Nico and Cserjan-Puschmann, Monika and Aguilar, Patricia Pereira and Auer, Simone and Demirtas, Halil and Andersson, Jakob and Lötsch, Felix and Holzer, Barbara and Steinrigl, Adi and Thalhammer, Florian and Schellnegger, Julia and Breuer, Monika and Knoll, Wolfgang and Strassl, Robert},
  issn         = {1873-4235},
  journal      = {Biosensors and Bioelectronics},
  publisher    = {Elsevier},
  title        = {{Bridging basic science and applied diagnostics: Comprehensive viral diagnostics enabled by graphene-based electronic biosensor technology advancements}},
  doi          = {10.1016/j.bios.2024.116807},
  volume       = {267},
  year         = {2025},
}

@misc{18967,
  abstract     = {Background: We identified small molecule tricyclic pyrone compound CP2 as a mild mitochondrial complex I (MCI) inhibitor that induces neuroprotection in multiple mouse models of AD. One of the major concerns while targeting mitochondria is the production of reactive oxygen species (ROS). CP2 consists of two diastereoisomers, D1 and D2, with distinct activity and toxicity profiles. This study was designed to understand how structure of D1 and D2 affects their binding to MCI and the consequential impact on ROS production.

Method: The X-ray crystallography and cryo-electron microscopy (cryo-EM) at global resolution of 3.25-3.27Å were employed to identify the molecular structure of D1 and D2 and the D1 binding to the isolated ovine MCI. The assessment of the MCI inhibition and the extent of ROS generation were done in isolated MCI and human neuroblastoma MC65 cells using flow cytometry, a Seahorse extracellular flux analyzer, and the kinetic studies.

Result: In the closed conformation of MCI, D1 selectively binds to the deep Quinone-site (Qd) but not to the shallow Q-site (Qs), sharing the same binding pocket as rotenone. In the open MCI state, D1 exclusively binds to the Qs in contrast to rotenone, which binds Qd and Qs in both closed and open states. At the same concentrations, D1 inhibits respiration to a greater extent compared to D2 (5:1 ratio) and produces higher level of ROS.

Conclusion:Cryo-EM unambiguously identified binding of D1 to both the Qd and Qs sites, contingent upon the conformational state of MCI. In contrast to rotenone, D1 binds Qd only in the closed conformation during catalytic cycle, leading to mild inhibition. Superimposing X-ray crystallography data of D1 and D2 onto cryo-EM data suggests that the orientation of the methyl group in D2 induces a flatter conformation, resulting in lower binding affinity to MCI, which correlates with lower inhibition and toxicity compared to D1. At physiologically relevant concentrations, CP2 (D1:D2 = 1:1) demonstrates low MCI inhibition yielding negligible ROS levels. This observation provides new insight into the absence of toxicity associated with CP2 treatment in vivo, further highlighting feasibility for the development of safe and efficacious MCI inhibitors.},
  author       = {Petrova, Olga and Trushin, Sergey A and Nguyen, Thi Kim Oanh and Ostroot, Mark and Schellenberg, Matthew and Johnson, Graham and Trushina, Eugenia and Sazanov, Leonid A},
  booktitle    = {Alzheimer's & Dementia},
  issn         = {1552-5279},
  number       = {S6},
  publisher    = {Wiley},
  title        = {{Structure‐activity relationship study of neuroprotective complex I inhibitor CP2}},
  doi          = {10.1002/alz.085971},
  volume       = {20},
  year         = {2024},
}

@article{15323,
  abstract     = {Supercomplexes of the respiratory chain are established constituents of the oxidative phosphorylation system, but their role in mammalian metabolism has been hotly debated. Although recent studies have shown that different tissues/organs are equipped with specific sets of supercomplexes, depending on their metabolic needs, the notion that supercomplexes have a role in the regulation of metabolism has been challenged. However, irrespective of the mechanistic conclusions, the composition of various high molecular weight supercomplexes remains uncertain. Here, using cryogenic electron microscopy, we demonstrate that mammalian (mouse) tissues contain three defined types of ‘respirasome’, supercomplexes made of CI, CIII2 and CIV. The stoichiometry and position of CIV differs in the three respirasomes, of which only one contains the supercomplex-associated factor SCAF1, whose involvement in respirasome formation has long been contended. Our structures confirm that the ‘canonical’ respirasome (the C-respirasome, CICIII2CIV) does not contain SCAF1, which is instead associated to a different respirasome (the CS-respirasome), containing a second copy of CIV. We also identify an alternative respirasome (A-respirasome), with CIV bound to the ‘back’ of CI, instead of the ‘toe’. This structural characterization of mouse mitochondrial supercomplexes allows us to hypothesize a mechanistic basis for their specific role in different metabolic conditions.},
  author       = {Vercellino, Irene and Sazanov, Leonid A},
  issn         = {1545-9985},
  journal      = {Nature Structural and Molecular Biology},
  pages        = {1061--1071},
  publisher    = {Springer Nature},
  title        = {{SCAF1 drives the compositional diversity of mammalian respirasomes}},
  doi          = {10.1038/s41594-024-01255-0},
  volume       = {31},
  year         = {2024},
}

@phdthesis{17319,
  abstract     = {This thesis comprises two distinct projects, each offering unique insights into fundamental
cellular processes. While distinct in their focus, these different perspectives have a common
theme: chemiosmotic theory and utilisation of the proton gradient for driving the essential
processes like auxin efflux and ATP synthesis, effectively bridging the membrane protein
structure and function from the realms of plant biology and cellular bioenergetics.
The first project of this thesis centres on the characterisation of PIN proteins, a class of
transmembrane transporters pivotal in the regulation of auxin transport and distribution in
plants. PINs form a conserved and phylogenetically abundant group of transporters present in
land plants and certain algae. Despite their great importance, they were one of the few elusive
proteins essential for plant development not to be structurally and mechanistically
characterised since their discovery almost 30 years ago. This work aimed to uncover the
structural and functional dynamics of the PIN protein-mediated auxin transport using an array
of experimental techniques, including protein purification, biochemical assays and structural
analysis. Through an exhaustive screening process that took several years and included testing
different PIN homologues, expression systems, constructs, and purification conditions, we
developed a robust protocol for isolating the pure, stable, and monodisperse PIN8 protein.
Moreover, utilising biophysical methods and buffer screening, we demonstrated that PIN8
exhibits detergent and pH-dependent stability, with mild detergents and lower pH (5.0 and 6.0)
being optimal for the stability of the protein. Using SEC-MALS and crosslinking, we
determined that PIN8 forms dimers, which was confirmed by our structural studies. We
obtained a cryo-EM map of PIN8 at pH 6.0, and, compared to recently published structures,
our map implies major pH-dependent conformational changes and possibly utilisation of the
proton gradient in the transport mechanism.
The subject of the second project was F1Fo-ATP synthase, an enzyme complex fundamental
to cellular energy metabolism. Through an approach integrating biochemical assays and
structural analysis, this research aimed to unveil the molecular mechanism of inhibition of ATP
synthase by yaku´amide, a bioactive compound with potential therapeutic implications. Using
submitochondrial particles and purified F1Fo-ATP synthase, we demonstrated that, contrary to
published data, yaku´amide inhibits both ATP hydrolysis and ATP synthesis reactions.
Moreover, we found that yaku´amide inhibitory activity is proton motive force (pmf)
dependent, with lower inhibition in a more coupled system. Utilising cryo-EM, we obtained
maps and models for the three main rotational states of murine ATP synthase (State 1 at 3.0 Å,
8
State 2 at 3.1 Å, and State 3 at 3.2 Å, overall). We observed several new features in our maps;
however, we cannot definitively determine the exact mechanism of yaku amide’s inhibition on
the protein due to either resolution limits or suboptimal binding of the inhibitor.},
  author       = {Lukic, Kristina},
  issn         = {2663-337X},
  pages        = {224},
  publisher    = {Institute of Science and Technology Austria},
  title        = {{Membrane proteins in plant physiology and bioenergetics : Investigating auxin efflux transporter PIN8 and ATP synthase inhibition by the novel inhibitor Yaku'amide B}},
  doi          = {10.15479/at:ista:17319},
  year         = {2024},
}

@article{12757,
  abstract     = {My group and myself have studied respiratory complex I for almost 30 years, starting in 1994 when it was known as a L-shaped giant ‘black box' of bioenergetics. First breakthrough was the X-ray structure of the peripheral arm, followed by structures of the membrane arm and finally the entire complex from Thermus thermophilus. The developments in cryo-EM technology allowed us to solve the first complete structure of the twice larger, ∼1 MDa mammalian enzyme in 2016. However, the mechanism coupling, over large distances, the transfer of two electrons to pumping of four protons across the membrane remained an enigma. Recently we have solved high-resolution structures of mammalian and bacterial complex I under a range of redox conditions, including catalytic turnover. This allowed us to propose a robust and universal mechanism for complex I and related protein families. Redox reactions initially drive conformational changes around the quinone cavity and a long-distance transfer of substrate protons. These set up a stage for a series of electrostatically driven proton transfers along the membrane arm (‘domino effect'), eventually resulting in proton expulsion from the distal antiporter-like subunit. The mechanism radically differs from previous suggestions, however, it naturally explains all the unusual structural features of complex I. In this review I discuss the state of knowledge on complex I, including the current most controversial issues.},
  author       = {Sazanov, Leonid A},
  issn         = {1470-8728},
  journal      = {The Biochemical Journal},
  number       = {5},
  pages        = {319--333},
  publisher    = {Portland Press},
  title        = {{From the 'black box' to 'domino effect' mechanism: What have we learned from the structures of respiratory complex I}},
  doi          = {10.1042/BCJ20210285},
  volume       = {480},
  year         = {2023},
}

@article{13232,
  abstract     = {The potential of immune-evasive mutation accumulation in the SARS-CoV-2 virus has led to its rapid spread, causing over 600 million confirmed cases and more than 6.5 million confirmed deaths. The huge demand for the rapid development and deployment of low-cost and effective vaccines against emerging variants has renewed interest in DNA vaccine technology. Here, we report the rapid generation and immunological evaluation of novel DNA vaccine candidates against the Wuhan-Hu-1 and Omicron variants based on the RBD protein fused with the Potato virus X coat protein (PVXCP). The delivery of DNA vaccines using electroporation in a two-dose regimen induced high-antibody titers and profound cellular responses in mice. The antibody titers induced against the Omicron variant of the vaccine were sufficient for effective protection against both Omicron and Wuhan-Hu-1 virus infections. The PVXCP protein in the vaccine construct shifted the immune response to the favorable Th1-like type and provided the oligomerization of RBD-PVXCP protein. Naked DNA delivery by needle-free injection allowed us to achieve antibody titers comparable with mRNA-LNP delivery in rabbits. These data identify the RBD-PVXCP DNA vaccine platform as a promising solution for robust and effective SARS-CoV-2 protection, supporting further translational study.},
  author       = {Dormeshkin, Dmitri and Katsin, Mikalai and Stegantseva, Maria and Golenchenko, Sergey and Shapira, Michail and Dubovik, Simon and Lutskovich, Dzmitry and Kavaleuski, Anton and Meleshko, Alexander},
  issn         = {2076-393X},
  journal      = {Vaccines},
  number       = {6},
  publisher    = {MDPI},
  title        = {{Design and immunogenicity of SARS-CoV-2 DNA vaccine encoding RBD-PVXCP fusion protein}},
  doi          = {10.3390/vaccines11061014},
  volume       = {11},
  year         = {2023},
}

@article{14040,
  abstract     = {Robust oxygenic photosynthesis requires a suite of accessory factors to ensure efficient assembly and repair of the oxygen-evolving photosystem two (PSII) complex. The highly conserved Ycf48 assembly factor binds to the newly synthesized D1 reaction center polypeptide and promotes the initial steps of PSII assembly, but its binding site is unclear. Here we use cryo-electron microscopy to determine the structure of a cyanobacterial PSII D1/D2 reaction center assembly complex with Ycf48 attached. Ycf48, a 7-bladed beta propeller, binds to the amino-acid residues of D1 that ultimately ligate the water-oxidising Mn4CaO5 cluster, thereby preventing the premature binding of Mn2+ and Ca2+ ions and protecting the site from damage. Interactions with D2 help explain how Ycf48 promotes assembly of the D1/D2 complex. Overall, our work provides valuable insights into the early stages of PSII assembly and the structural changes that create the binding site for the Mn4CaO5 cluster.},
  author       = {Zhao, Ziyu and Vercellino, Irene and Knoppová, Jana and Sobotka, Roman and Murray, James W. and Nixon, Peter J. and Sazanov, Leonid A and Komenda, Josef},
  issn         = {2041-1723},
  journal      = {Nature Communications},
  publisher    = {Springer Nature},
  title        = {{The Ycf48 accessory factor occupies the site of the oxygen-evolving manganese cluster during photosystem II biogenesis}},
  doi          = {10.1038/s41467-023-40388-6},
  volume       = {14},
  year         = {2023},
}

@phdthesis{12781,
  abstract     = {Most energy in humans is produced in form of ATP by the mitochondrial respiratory chain consisting of several protein assemblies embedded into lipid membrane (complexes I-V). Complex I is the first and the largest enzyme of the respiratory chain which is essential for energy production. It couples the transfer of two electrons from NADH to ubiquinone with proton translocation across bacterial or inner mitochondrial membrane. The coupling mechanism between electron transfer and proton translocation is one of the biggest enigma in bioenergetics and structural biology. Even though the enzyme has been studied for decades, only recent technological advances in cryo-EM allowed its extensive structural investigation. 

Complex I from E.coli appears to be of special importance because it is a perfect model system with a rich mutant library, however the structure of the entire complex was unknown. In this thesis I have resolved structures of the minimal complex I version from E. coli in different states including reduced, inhibited, under reaction turnover and several others. Extensive structural analyses of these structures and comparison to structures from other species allowed to derive general features of conformational dynamics and propose a universal coupling mechanism. The mechanism is straightforward, robust and consistent with decades of experimental data available for complex I from different species. 

Cyanobacterial NDH (cyanobacterial complex I) is a part of broad complex I superfamily and was studied as well in this thesis. It plays an important role in cyclic electron transfer (CET), during which electrons are cycled within PSI through ferredoxin and plastoquinone to generate proton gradient without NADPH production. Here, I solved structure of NDH and revealed additional state, which was not observed before. The novel “resting” state allowed to propose the mechanism of CET regulation. Moreover, conformational dynamics of NDH resembles one in complex I which suggest more broad universality of the proposed coupling mechanism.

In summary, results presented here helped to interpret decades of experimental data for complex I and contributed to fundamental mechanistic understanding of protein function.
},
  author       = {Kravchuk, Vladyslav},
  isbn         = {978-3-99078-029-9},
  issn         = {2663-337X},
  pages        = {127},
  publisher    = {Institute of Science and Technology Austria},
  title        = {{Structural and mechanistic study of bacterial complex I and its cyanobacterial ortholog}},
  doi          = {10.15479/at:ista:12781},
  year         = {2023},
}

@article{10945,
  abstract     = {Mica-titania pearlescent pigments (MTs) were previously coated with organic molecules to obtain combination pigments (CPs) for achieving certain improvements or functionalities. Anthocyanins (ACNs) are molecules that can be extracted from natural resources and exhibit color changes via pH modifications of the enclosing medium. The purpose of the study was to produce a new series of CPs by depositing ACNs on MTs at different pH values, to observe the changes in color, and to associate these changes to thermogravimetrically determined deposition efficiencies in light of spectral differences. The extraction and deposition methods were based on aqueous chemistry and were straightforward. The ACN deposition generally increased with increasing pH and correlated with the consistency between the charges of the MT surfaces and the dominant ACN species at a specific pH value. The fluorescence of the CPs was inversely correlated with the deposition quantities invoking the possibility of a quenching effect.},
  author       = {Çoruh, Mehmet Orkun and Gündüz, Güngör and Çolak, Üner and Maviş, Bora},
  issn         = {2079-6447},
  journal      = {Colorants},
  number       = {2},
  pages        = {149--164},
  publisher    = {MDPI},
  title        = {{pH-dependent coloring of combination effect pigments with anthocyanins from Brassica oleracea var. capitata F. rubra}},
  doi          = {10.3390/colorants1020010},
  volume       = {1},
  year         = {2022},
}

@article{11167,
  abstract     = {Complex I is one of the major respiratory complexes, conserved from bacteria to mammals. It oxidises NADH, reduces quinone and pumps protons across the membrane, thus playing a central role in the oxidative energy metabolism. In this review we discuss our current state of understanding the structure of complex I from various species of mammals, plants, fungi, and bacteria, as well as of several complex I-related proteins. By comparing the structural evidence from these systems in different redox states and data from mutagenesis and molecular simulations, we formulate the mechanisms of electron transfer and proton pumping and explain how they are conformationally and electrostatically coupled. Finally, we discuss the structural basis of the deactivation phenomenon in mammalian complex I.},
  author       = {Kampjut, Domen and Sazanov, Leonid A},
  issn         = {0959-440X},
  journal      = {Current Opinion in Structural Biology},
  keywords     = {Molecular Biology, Structural Biology},
  publisher    = {Elsevier},
  title        = {{Structure of respiratory complex I – An emerging blueprint for the mechanism}},
  doi          = {10.1016/j.sbi.2022.102350},
  volume       = {74},
  year         = {2022},
}

@article{11462,
  abstract     = {Nanobodies (VHH) from camelid antibody libraries hold great promise as therapeutic agents and components of immunoassay systems. Synthetic antibody libraries that could be designed and generated once and for various applications could yield binders to virtually any targets, even for non-immunogenic or toxic ones, in a short term. One of the most difficult tasks is to obtain antibodies with a high affinity and specificity to polyglycosylated proteins. It requires antibody libraries with extremely high functional diversity and the use of sophisticated selection techniques. Here we report a development of a novel sandwich immunoassay involving a combination of the synthetic library-derived VHH-Fc fusion protein as a capture antibody and the immune single-chain fragment variable (scFv) as a tracer for the detection of pregnancy-associated glycoprotein (PAG) of cattle (Bos taurus). We succeeded in the generation of a number of specific scFv antibodies against PAG from the mouse immune library. Subsequent selection using the immobilized scFv-Fc capture antibody allowed to isolate 1.9 nM VHH binder from the diverse synthetic library without any overlapping with the capture antibody binding site. The prototype sandwich ELISA based on the synthetic VHH and the immune scFv was established. This is the first successful example of the combination of synthetic and immune antibody libraries in a single sandwich immunoassay. Thus, our approach could be used for the express isolation of antibody pairs and the development of sandwich immunoassays for challenging antigens.},
  author       = {Dormeshkin, Dmitri and Shapira, Michail and Karputs, Alena and Kavaleuski, Anton and Kuzminski, Ivan and Stepanova, Elena and Gilep, Andrei},
  issn         = {1432-0614},
  journal      = {Applied Microbiology and Biotechnology},
  pages        = {5093--5103},
  publisher    = {Springer Nature},
  title        = {{Combining of synthetic VHH and immune scFv libraries for pregnancy-associated glycoproteins ELISA development}},
  doi          = {10.1007/s00253-022-12022-w},
  volume       = {106},
  year         = {2022},
}

@article{11551,
  abstract     = {Imbalanced mitochondrial dNTP pools are known players in the pathogenesis of multiple human diseases. Here we show that, even under physiological conditions, dGTP is largely overrepresented among other dNTPs in mitochondria of mouse tissues and human cultured cells. In addition, a vast majority of mitochondrial dGTP is tightly bound to NDUFA10, an accessory subunit of complex I of the mitochondrial respiratory chain. NDUFA10 shares a deoxyribonucleoside kinase (dNK) domain with deoxyribonucleoside kinases in the nucleotide salvage pathway, though no specific function beyond stabilizing the complex I holoenzyme has been described for this subunit. We mutated the dNK domain of NDUFA10 in human HEK-293T cells while preserving complex I assembly and activity. The NDUFA10E160A/R161A shows reduced dGTP binding capacity in vitro and leads to a 50% reduction in mitochondrial dGTP content, proving that most dGTP is directly bound to the dNK domain of NDUFA10. This interaction may represent a hitherto unknown mechanism regulating mitochondrial dNTP availability and linking oxidative metabolism to DNA maintenance.},
  author       = {Molina-Granada, David and González-Vioque, Emiliano and Dibley, Marris G. and Cabrera-Pérez, Raquel and Vallbona-Garcia, Antoni and Torres-Torronteras, Javier and Sazanov, Leonid A and Ryan, Michael T. and Cámara, Yolanda and Martí, Ramon},
  issn         = {2399-3642},
  journal      = {Communications Biology},
  number       = {1},
  publisher    = {Springer Nature},
  title        = {{Most mitochondrial dGTP is tightly bound to respiratory complex I through the NDUFA10 subunit}},
  doi          = {10.1038/s42003-022-03568-6},
  volume       = {5},
  year         = {2022},
}

@article{11648,
  abstract     = {Progress in structural membrane biology has been significantly accelerated by the ongoing 'Resolution Revolution' in cryo electron microscopy (cryo-EM). In particular, structure determination by single particle analysis has evolved into the most powerful method for atomic model building of multisubunit membrane protein complexes. This has created an ever increasing demand in cryo-EM machine time, which to satisfy is in need of new and affordable cryo electron microscopes. Here, we review our experience in using the JEOL CRYO ARM 200 prototype for the structure determination by single particle analysis of three different multisubunit membrane complexes: the Thermus thermophilus V-type ATPase VO complex, the Thermosynechococcus elongatus photosystem I monomer and the flagellar motor LP-ring from Salmonella enterica.},
  author       = {Gerle, Christoph and Kishikawa, Jun-ichi and Yamaguchi, Tomoko and Nakanishi, Atsuko and Çoruh, Mehmet Orkun and Makino, Fumiaki and Miyata, Tomoko and Kawamoto, Akihiro and Yokoyama, Ken and Namba, Keiichi and Kurisu, Genji and Kato, Takayuki},
  issn         = {2050-5701},
  journal      = {Microscopy},
  keywords     = {Radiology, Nuclear Medicine and imaging, Instrumentation, Structural Biology},
  number       = {5},
  pages        = {249--261},
  publisher    = {Oxford University Press},
  title        = {{Structures of multisubunit membrane complexes with the CRYO ARM 200}},
  doi          = {10.1093/jmicro/dfac037},
  volume       = {71},
  year         = {2022},
}

@article{12252,
  abstract     = {The COVID−19 pandemic not only resulted in a global crisis, but also accelerated vaccine development and antibody discovery. Herein we report a synthetic humanized VHH library development pipeline for nanomolar-range affinity VHH binders to SARS-CoV-2 variants of concern (VoC) receptor binding domains (RBD) isolation. Trinucleotide-based randomization of CDRs by Kunkel mutagenesis with the subsequent rolling-cycle amplification resulted in more than 10<jats:sup>11</jats:sup> diverse phage display library in a manageable for a single person number of electroporation reactions. We identified a number of nanomolar-range affinity VHH binders to SARS-CoV-2 variants of concern (VoC) receptor binding domains (RBD) by screening a novel synthetic humanized antibody library. In order to explore the most robust and fast method for affinity improvement, we performed affinity maturation by CDR1 and CDR2 shuffling and avidity engineering by multivalent trimeric VHH fusion protein construction. As a result, H7-Fc and G12x3-Fc binders were developed with the affinities in nM and pM range respectively. Importantly, these affinities are weakly influenced by most of SARS-CoV-2 VoC mutations and they retain moderate binding to BA.4\5. The plaque reduction neutralization test (PRNT) resulted in IC50 = 100 ng\ml and 9.6 ng\ml for H7-Fc and G12x3-Fc antibodies, respectively, for the emerging Omicron BA.1 variant. Therefore, these VHH could expand the present landscape of SARS-CoV-2 neutralization binders with the therapeutic potential for present and future SARS-CoV-2 variants.},
  author       = {Dormeshkin, Dmitri and Shapira, Michail and Dubovik, Simon and Kavaleuski, Anton and Katsin, Mikalai and Migas, Alexandr and Meleshko, Alexander and Semyonov, Sergei},
  issn         = {1664-3224},
  journal      = {Frontiers in Immunology},
  keywords     = {Immunology, Immunology and Allergy, COVID-19, SARS-CoV-2, synthetic library, RBD, neutralization nanobody, VHH},
  publisher    = {Frontiers Media},
  title        = {{Isolation of an escape-resistant SARS-CoV-2 neutralizing nanobody from a novel synthetic nanobody library}},
  doi          = {10.3389/fimmu.2022.965446},
  volume       = {13},
  year         = {2022},
}

@article{12282,
  abstract     = {From a simple thought to a multicellular movement},
  author       = {Amberg, Nicole and Stouffer, Melissa A and Vercellino, Irene},
  issn         = {1477-9137},
  journal      = {Journal of Cell Science},
  number       = {8},
  publisher    = {The Company of Biologists},
  title        = {{Operation STEM fatale – how an equity, diversity and inclusion initiative has brought us to reflect on the current challenges in cell biology and science as a whole}},
  doi          = {10.1242/jcs.260017},
  volume       = {135},
  year         = {2022},
}