Journal of structural biology最新文献_第2页

Improving CryoEM maps of symmetry-mismatched macromolecular assemblies: A case study on the flagellar motor

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-03-05 DOI: 10.1016/j.jsb.2025.108184

Prashant K. Singh , T.M. Iverson

{"title":"Improving CryoEM maps of symmetry-mismatched macromolecular assemblies: A case study on the flagellar motor","authors":"Prashant K. Singh , T.M. Iverson","doi":"10.1016/j.jsb.2025.108184","DOIUrl":"10.1016/j.jsb.2025.108184","url":null,"abstract":"<div><div>Advances in cryo-electron microscopy instrumentation and sample preparation have significantly improved the ability to collect quality data for biomolecular structures. However, achieving resolutions consistent with data quality remains challenging in structures with symmetry mismatches. As a case study, the bacterial flagellar motor is a large complex essential for bacterial chemotaxis and virulence. This motor contains a smaller membrane-supramembrane ring (MS-ring) and a larger cytoplasmic ring (C-ring). These two features have a 33:34 symmetry mismatch when expressed in <em>E. coli</em>. Because close symmetry mismatches are the most difficult to deconvolute, this makes the flagellar motor an excellent model system to evaluate refinement strategies for symmetry mismatch. We compared the performance of masked refinement, local refinement, and particle subtracted refinement on the same data. We found that particle subtraction prior to refinement was critical for approaching the smaller MS-ring. Additional processing resulted in final resolutions of 3.1 Å for the MS-ring and 3.0 Å for the C-ring, which improves the resolution of the MS-ring by 0.3 Å and the resolution of the C-ring by 1.0 Å as compared to past work. Although particle subtraction is fairly well-established, it is rarely applied to problems of symmetry mismatch, making this case study a valuable demonstration of its utility in this context.</div></div>","PeriodicalId":17074,"journal":{"name":"Journal of structural biology","volume":"217 2","pages":"Article 108184"},"PeriodicalIF":3.0,"publicationDate":"2025-03-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143585985","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Cryo-iCLEM: Cryo correlative light and electron microscopy with immersion objectives

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-03-01 DOI: 10.1016/j.jsb.2025.108179

Niko Faul , Shih-Ya Chen , Christian Lamberz , Mark Bruckner , Christian Dienemann , Thomas P. Burg

{"title":"Cryo-iCLEM: Cryo correlative light and electron microscopy with immersion objectives","authors":"Niko Faul , Shih-Ya Chen , Christian Lamberz , Mark Bruckner , Christian Dienemann , Thomas P. Burg","doi":"10.1016/j.jsb.2025.108179","DOIUrl":"10.1016/j.jsb.2025.108179","url":null,"abstract":"<div><div>Correlative light and electron microscopy (CLEM) is a powerful tool for investigating cellular structure and function at the molecular level. However, while electron microscopy is often performed to great advantage at cryogenic temperatures, this is not always the case for light microscopy. One key challenge is the lack of cryo-compatible immersion objectives. In recent years, multiple cryoimmersion light microscopy (cryo-iLM) approaches have been described, but these techniques have never been used in correlative approaches. Here we present a novel workflow for correlative cryoimmersion light microscopy and electron cryomicroscopy (cryo-iCLEM). Cryo-electron tomography conducted before and after cryo-iLM reveals that cryo-iCLEM maintains ultra-thin, electron-transparent samples mechanically intact and does not degrade the ultrastructural preservation achieved through plunge-freezing. For cryo-iLM, the sample is first embedded in a viscous immersion medium at cryogenic temperatures and examined with a custom cryo-immersion objective. After cryo-iLM, the immersion medium is dissolved in liquid ethane, allowing for subsequent cryo-EM imaging. We further show that cryo-iCLEM can be used on FIB-lamellae, demonstrating that mechanically sensitive samples remain undamaged. Embedding the sample in the immersion fluid reduces contamination and thus allows data acquisition over many hours. Samples can therefore be examined in detail with the advantage of low bleaching rates of fluorophores at cryogenic temperatures. In the future, we hope that our approach can help improve the performance of many advanced light microscopy techniques when they are applied in the context of cryo-CLEM.</div></div>","PeriodicalId":17074,"journal":{"name":"Journal of structural biology","volume":"217 1","pages":"Article 108179"},"PeriodicalIF":3.0,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143458330","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Computational Engineering of siderocalin to modulate its binding affinity to the antihypertension drug candesartan

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-03-01 DOI: 10.1016/j.jsb.2025.108180

Kristina Vogel , Johanna Moeller , Nina G. Bozhanova , Markus Voehler , Anja Penk , Jens Meiler , Clara T. Schoeder

{"title":"Computational Engineering of siderocalin to modulate its binding affinity to the antihypertension drug candesartan","authors":"Kristina Vogel , Johanna Moeller , Nina G. Bozhanova , Markus Voehler , Anja Penk , Jens Meiler , Clara T. Schoeder","doi":"10.1016/j.jsb.2025.108180","DOIUrl":"10.1016/j.jsb.2025.108180","url":null,"abstract":"<div><div>Lipocalin family proteins have been shown to bind a vast array of small molecules and have subsequently been adapted to selectively bind specific ligands. In this study, candesartan, an antihypertension drug, was identified to bind mouse and human siderocalin in biomolecular NMR experiments, allowing for structural insights into the candesartan-siderocalin interaction. The ligand binding site was determined through an integrative structural biology approach using <em>in silico</em> ligand docking guided by NMR experiments. Building on this structurally informed binding model, we used rational protein design to modulate the binding pocket for increased or decreased ligand binding affinity. The predicted mutations were evaluated <em>in vitro</em> using isothermal titration calorimetry. This resulted in a mutant with a 50-fold increase in binding affinity in addition to a second mutant with a five-fold decrease in binding affinity. Thus, siderocalins have potential as a scaffold for creation of various ligand binding-based tools, including drug scavengers.</div></div>","PeriodicalId":17074,"journal":{"name":"Journal of structural biology","volume":"217 1","pages":"Article 108180"},"PeriodicalIF":3.0,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143468251","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Lack of embryonic skeletal muscle in mice leads to abnormal mineral deposition and growth

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-03-01 DOI: 10.1016/j.jsb.2025.108178

Isabella Silva Barreto , Marianne Liebi , Sophie Le Cann , Saima Ahmed , Leonard C. Nielsen , Tilman A. Grünewald , Hector Dejea , Viviane Lutz-Bueno , Niamh C. Nowlan , Hanna Isaksson

{"title":"Lack of embryonic skeletal muscle in mice leads to abnormal mineral deposition and growth","authors":"Isabella Silva Barreto , Marianne Liebi , Sophie Le Cann , Saima Ahmed , Leonard C. Nielsen , Tilman A. Grünewald , Hector Dejea , Viviane Lutz-Bueno , Niamh C. Nowlan , Hanna Isaksson","doi":"10.1016/j.jsb.2025.108178","DOIUrl":"10.1016/j.jsb.2025.108178","url":null,"abstract":"<div><div>Developing bones can be severely impaired by a range of disorders where muscular loading is abnormal. Recent work has indicated that the effects of absent skeletal muscle on bones are more severe early in development, with rudiment length and mineralization lengths being almost normal in muscle-less limbs just prior to birth. However, the impact of abnormal mechanical loading on the nanoscale structure and composition during prenatal mineralization remains unknown. In this exploratory study, we characterized the mineralization process of humeri from muscle-less limb embryonic mice using a multiscale approach by combining X-ray scattering and fluorescence with infrared and light microscopy to identify potential key aspects of interest for future in-depth investigations. Muscle-less humeri were characterized by initially less mineralized tissue to later catch up with controls, and exhibited continuous growth of mineral particles, which ultimately led to seemingly larger mineral particles than their controls at the end of development. Muscle-less limbs exhibited an abnormal pattern of mineralization, reflected by a more widespread distribution of zinc and homogenous distribution of hydroxyapatite compared to controls, which instead showed trabecular-like structures and zinc localized only to regions of ongoing mineralization. The decrease in collagen content in the hypertrophic zone due to resorption of the cartilage collagen matrix was less distinct in muscle-less limbs compared to controls. Surprisingly, the nanoscale orientation of the mineral particles was unaffected by the lack of skeletal muscle. The identified accelerated progression of ossification in muscle-less limbs at later prenatal stages provides a possible anatomical mechanism underlying their recovery in skeletal development.</div></div>","PeriodicalId":17074,"journal":{"name":"Journal of structural biology","volume":"217 1","pages":"Article 108178"},"PeriodicalIF":3.0,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143433421","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Effects of Gα C-terminal deletion on the intrinsic GDP release/GTPase activity and conformational dynamics Gα C 端缺失对固有 GDP 释放/GTP 酶活性和构象动态的影响

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-02-27 DOI: 10.1016/j.jsb.2025.108182

Junyoung Kim, Ka Young Chung

{"title":"Effects of Gα C-terminal deletion on the intrinsic GDP release/GTPase activity and conformational dynamics","authors":"Junyoung Kim, Ka Young Chung","doi":"10.1016/j.jsb.2025.108182","DOIUrl":"10.1016/j.jsb.2025.108182","url":null,"abstract":"<div><div>Heterotrimeric G proteins (G proteins) serve as key signaling mediators downstream of G protein-coupled receptors (GPCRs). Comprised of Gα, Gβ, and Gγ subunits, the activation state of Gα, determined by GDP or GTP binding, governs G protein activity. While high-resolution structures of GPCR-G protein complexes have identified the Gα C-terminal 5 residues (<em>i.e.,</em> wavy hook) as critical for GPCR binding and coupling selectivity, its influence on Gα’s intrinsic biochemical properties remains unclear. Here, we investigated the role of wavy hook truncation in the intrinsic GDP/GTP turnover rate, GTPase activity, and conformational dynamics of Gαs and Gαi1 using BODIPY-labeled nucleotides and hydrogen/deuterium exchange mass spectrometry (HDX-MS). Truncation of the wavy hook significantly altered the GDP/GTP turnover rate, GTPase activity, and conformational flexibility of Gαs, particularly at the p-loop through α1 region, but had minimal impact on Gαi1. These findings reveal subtype-specific effects of the wavy hook on G protein stability and conformational dynamics, highlighting the importance of structural elements in regulating G protein function and their implications for GPCR signaling studies.</div></div>","PeriodicalId":17074,"journal":{"name":"Journal of structural biology","volume":"217 2","pages":"Article 108182"},"PeriodicalIF":3.0,"publicationDate":"2025-02-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143537316","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Three-dimensional reconstruction of Magnetofaba australis strain IT-1: Magnetosome chain position with respect to flagella

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-02-26 DOI: 10.1016/j.jsb.2025.108181

Eduardo Monteiro , Anderson S. Cabral , Viviana Morillo , Daniel Acosta-Avalos , Ulysses Lins , Fernanda Abreu

{"title":"Three-dimensional reconstruction of Magnetofaba australis strain IT-1: Magnetosome chain position with respect to flagella","authors":"Eduardo Monteiro , Anderson S. Cabral , Viviana Morillo , Daniel Acosta-Avalos , Ulysses Lins , Fernanda Abreu","doi":"10.1016/j.jsb.2025.108181","DOIUrl":"10.1016/j.jsb.2025.108181","url":null,"abstract":"<div><div>Magnetotactic bacteria (MTB) are a broad and diverse group of Gram-negative prokaryotes that biomineralize magnetosomes, organelles composed of a magnetic nanocrystal of magnetite (Fe<sub>3</sub>O<sub>4</sub>) or greigite (Fe<sub>3</sub>S<sub>4</sub>) and enveloped by a biological membrane. Magnetosomes are arranged in one or more chains intracellularly, which impart a magnetic moment to the cell. These structures permit a passive orientation of the MTB with the geomagnetic field lines (GML), which, when associated with swimming propelled by flagella, originate a phenomenon called magneto-aerotaxis, an important life strategy in a chemical stratified environment. There is a classical model based on elongated cells as vibrios and rods that tries to explain the magneto-aerotaxis. Still, this model raises questions when applied to other morphologies other than elongated cells. Here, we observe the spatial disposition of magnetosomes, motility behavior, and influence of magneto-aerotaxis in <em>Magnetofaba australis</em> strain IT-1, an MTB that achieves high swimming speeds and has some peculiarity in its motility. The three-dimensional reconstruction showed that <em>Mf. australis</em> strain IT-1′s magnetosome chain is misaligned with the swimming axis, which makes it impossible to use the classical model to explain magneto-aerotaxis in this MTB. Despite this, <em>Mf. australis</em> strain IT-1 was capable of swimming aligned to the GML. Also, this work studied the influence of the magnetosome and magneto-aerotaxis between populations of <em>Mf. australis</em> strain IT-1 with and without magnetosomes. Our results indicated that the magnetosome presence not only positively influences the movement in <em>Mf.australis</em> strain IT-1 but also can positively impact population growth in these MTB.</div></div>","PeriodicalId":17074,"journal":{"name":"Journal of structural biology","volume":"217 2","pages":"Article 108181"},"PeriodicalIF":3.0,"publicationDate":"2025-02-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143531298","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Computational identification of B and T-cell epitopes for designing a multi-epitope vaccine against SARS-CoV-2 spike glycoprotein

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-02-11 DOI: 10.1016/j.jsb.2025.108177

Truc Ly Nguyen , Thong Ba Nguyen , Heebal Kim

{"title":"Computational identification of B and T-cell epitopes for designing a multi-epitope vaccine against SARS-CoV-2 spike glycoprotein","authors":"Truc Ly Nguyen , Thong Ba Nguyen , Heebal Kim","doi":"10.1016/j.jsb.2025.108177","DOIUrl":"10.1016/j.jsb.2025.108177","url":null,"abstract":"<div><div>Although the peak of the COVID-19 pandemic has passed, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to pose a significant global threat and remains a public health concern. Given the ongoing risk and the substantial loss of life caused by the virus, continuous research into vaccine development is essential. This study employs immunoinformatics approaches to identify T-cell and B-cell epitopes for designing a multi-epitope peptide vaccine candidate targeting the Omicron variant. The proposed vaccine construct comprises 1435 amino acids, including eight linear B lymphocyte, seven cytotoxic T lymphocyte, and five helper T lymphocyte epitopes, along with appropriate adjuvants and linkers. The evaluation of the vaccine revealed high antigenicity, non-allergenicity, non-toxicity, and favorable physicochemical properties. To further assess its efficacy, molecular docking studies were performed to investigate interactions between the vaccine and key immune components, including Toll-like receptors and major histocompatibility complex molecules. Stability of these interactions was confirmed using molecular dynamics simulations in triplicate, conducted over 100 ns using GROMACS 2023 to compute key metrics, such as root mean square deviation, root mean square fluctuation, solvent-accessible surface area, and radius of gyration. The results demonstrate that the multi-epitope vaccine has the potential to elicit strong immune responses against the Omicron variant, providing a promising foundation for further experimental validation and clinical development in COVID-19 vaccine research.</div></div>","PeriodicalId":17074,"journal":{"name":"Journal of structural biology","volume":"217 2","pages":"Article 108177"},"PeriodicalIF":3.0,"publicationDate":"2025-02-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143414501","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A computational approach to predict the effects of missense mutations on protein amyloidogenicity: A case study in hereditary transthyretin cardiomyopathy

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-02-09 DOI: 10.1016/j.jsb.2025.108176

Ivan A. Pyankov , Valentin Gonay , Yaroslav A. Stepanov , Pavel Shestun , Anna A. Kostareva , Mayya V. Uspenskaya , Michael G. Petukhov , Andrey V. Kajava

{"title":"A computational approach to predict the effects of missense mutations on protein amyloidogenicity: A case study in hereditary transthyretin cardiomyopathy","authors":"Ivan A. Pyankov , Valentin Gonay , Yaroslav A. Stepanov , Pavel Shestun , Anna A. Kostareva , Mayya V. Uspenskaya , Michael G. Petukhov , Andrey V. Kajava","doi":"10.1016/j.jsb.2025.108176","DOIUrl":"10.1016/j.jsb.2025.108176","url":null,"abstract":"<div><div>With many amyloidosis-associated missense mutations still unidentified and early diagnostic methods largely unavailable, there is an urgent need for a reliable computational approach to predict hereditary amyloidoses from gene sequencing data. Progress has been made in predicting amyloidosis-triggering sequences within intrinsically disordered regions. However, some diseases are caused by mutations in amyloidogenic regions within structured domains that must unfold for amyloid formation. Accurate prediction of amyloidogenic regions requires tools for detecting amyloidogenicity and assessing mutation effects on protein stability. We developed datasets of mutations linked to hereditary ATTR cardiomyopathy and others likely unrelated, evaluating TTR mutants with amyloidogenicity and stability predictors. Notably, the stability predictors consistently indicated that ATTR-related mutations tend to destabilize the TTR structure more than non-ATTR-associated mutations. Using these datasets and newly generated mutation features, we developed a machine learning model SDAM-TTR to predict mutations leading to ATTR cardiomyopathy.</div></div>","PeriodicalId":17074,"journal":{"name":"Journal of structural biology","volume":"217 1","pages":"Article 108176"},"PeriodicalIF":3.0,"publicationDate":"2025-02-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143395698","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Interaction of berberine with different forms of DNA in human telomeric region

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-02-01 DOI: 10.1016/j.jsb.2025.108175

Mahdiyeh Mostafavi , Leila Hassani , Maryam Khoshkam

{"title":"Interaction of berberine with different forms of DNA in human telomeric region","authors":"Mahdiyeh Mostafavi , Leila Hassani , Maryam Khoshkam","doi":"10.1016/j.jsb.2025.108175","DOIUrl":"10.1016/j.jsb.2025.108175","url":null,"abstract":"<div><div>Guanine-rich oligonucleotide sequences have the potential to form four-stranded structure known as G-quadruplex. These structures are frequently observed in crucial regions of the human genome, including promoter and telomeric regions. Due to their involvement in regulating gene expression and cell division, G-quadruplexes have emerged as promising targets for anticancer drugs. This study investigated interaction of berberine with different forms of DNA within human telomeric region. The results of absorption and fluorescence spectroscopy indicated that conformation of DNA plays an important role in the mode of binding. Circular dichroism suggested that berberine promotes compaction of the unstable quadruplex structure formed under non-saline conditions. Furthermore, interaction of berberine with the stable structures of G-quadruplex resulted in a change in their compactness without altering the type of DNA structure. 3D fluorescence spectra analysis by chemometrics methods showed formation of two distinct species probably attributed to the self-association and specific binding of berberin to the different forms of DNA. It can be also concluded that berberine forms a more stable complex with the human telomeric hybride type G-quadruplex structure compared with the basket type. In conclusion, the findings imply that the successful design of drugs targeting DNA within the human telomere region necessitates careful consideration of the diverse forms of DNA.</div></div>","PeriodicalId":17074,"journal":{"name":"Journal of structural biology","volume":"217 1","pages":"Article 108175"},"PeriodicalIF":3.0,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143122992","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Different strategies towards strength: Unveiling the role of Zn vs Mn/Ca and chitin arrangement in scorpion stingers

IF 3 3区生物学

Journal of structural biology Pub Date : 2025-01-30 DOI: 10.1016/j.jsb.2025.108174

C. Sakr , P. Cook , M. Seiter , C. Hörweg , S. Žák , M. Cordill , M. Burghammer , M. Sztucki , H. Lichtenegger

引用次数: 0