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Dr. Przemyslaw Grela
Maria Curie-Sklodowska University, Department of Molecular Biology

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0 Ribosome inactivating proteins

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Research article
Published: 30 July 2021 in FEBS Letters
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The ribosome is subjected to post-translational modifications, including phosphorylation, that affect its biological activity. Among ribosomal elements, the P-proteins undergo phosphorylation within the C-terminus, the element which interacts with trGTPases or ribosome inactivating proteins (RIPs), however the role of phosphorylation has never been elucidated. Here, we probed the function of phosphorylation on the interaction of P-proteins with RIPs using the ribosomal P1-P2 dimer. We determined the kinetic parameters of the interaction with the toxins using biolayer interferometry and microscale thermophoresis. The results present the first mechanistic insight into the function of P-protein phosphorylation, showing that introduction of a negative charge into the C-terminus of P1-P2 proteins promotes α-helix formation and decreases the affinity of the P-proteins for the RIPs.

ACS Style

Patrycja Horbowicz‐Drożdżal; Karol Kamel; Sebastian Kmiecik; Lidia Borkiewicz; Nilgun E. Tumer; Pang‐Chui Shaw; Marek Tchórzewski; Przemysław Grela. Phosphorylation of the conserved C‐terminal domain of ribosomal P‐proteins impairs the mode of interaction with plant toxins. FEBS Letters 2021, 1 .

AMA Style

Patrycja Horbowicz‐Drożdżal, Karol Kamel, Sebastian Kmiecik, Lidia Borkiewicz, Nilgun E. Tumer, Pang‐Chui Shaw, Marek Tchórzewski, Przemysław Grela. Phosphorylation of the conserved C‐terminal domain of ribosomal P‐proteins impairs the mode of interaction with plant toxins. FEBS Letters. 2021; ():1.

Chicago/Turabian Style

Patrycja Horbowicz‐Drożdżal; Karol Kamel; Sebastian Kmiecik; Lidia Borkiewicz; Nilgun E. Tumer; Pang‐Chui Shaw; Marek Tchórzewski; Przemysław Grela. 2021. "Phosphorylation of the conserved C‐terminal domain of ribosomal P‐proteins impairs the mode of interaction with plant toxins." FEBS Letters , no. : 1.

Journal article
Published: 22 March 2021 in Biomolecules
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A white rot fungus Cerrena unicolor has been identified as an important source of laccase, unfortunately regulation of this enzyme genes expression is poorly understood. Using 1D and 2D PAGE and LC-MS/MS, laccase isoenzymes were investigated in the liquid filtrate of C. unicolor culture. The level of expression of laccase genes was measured using qPCR. The elevated concentrations of copper and manganese in the medium caused greatest change in genes expression and three laccase transcripts were significantly affected after culture temperature was decreased from 28 to 4 °C or increased to 40 °C. The small differences in the PAGE band intensities of individual laccase proteins were also observed, indicating that given compound affect particular laccase’s transcript. Analyses of laccase-specific activity, at all tested conditions, showed the increased activities as compared to the control, suggesting that enzyme is regulated at the post-translational stage. We observed that the aspartic protease purified from C. unicolor, significantly stimulate laccase activity. Moreover, electrochemical analysis of protease-treated laccase sample had 5 times higher redox peaks. The obtained results indicate that laccases released by C. unicolor are regulated at transcriptional, translational, and at the post-translational steps of gene expression helping fungus adapt to the environmental changes.

ACS Style

Anna Pawlik; Beata Ciołek; Justyna Sulej; Andrzej Mazur; Przemysław Grela; Magdalena Staszczak; Mateusz Niścior; Magdalena Jaszek; Anna Matuszewska; Grzegorz Janusz; Andrzej Paszczyński. Cerrena unicolor Laccases, Genes Expression and Regulation of Activity. Biomolecules 2021, 11, 468 .

AMA Style

Anna Pawlik, Beata Ciołek, Justyna Sulej, Andrzej Mazur, Przemysław Grela, Magdalena Staszczak, Mateusz Niścior, Magdalena Jaszek, Anna Matuszewska, Grzegorz Janusz, Andrzej Paszczyński. Cerrena unicolor Laccases, Genes Expression and Regulation of Activity. Biomolecules. 2021; 11 (3):468.

Chicago/Turabian Style

Anna Pawlik; Beata Ciołek; Justyna Sulej; Andrzej Mazur; Przemysław Grela; Magdalena Staszczak; Mateusz Niścior; Magdalena Jaszek; Anna Matuszewska; Grzegorz Janusz; Andrzej Paszczyński. 2021. "Cerrena unicolor Laccases, Genes Expression and Regulation of Activity." Biomolecules 11, no. 3: 468.

Journal article
Published: 21 July 2020 in Cells
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Aging is a biological phenomenon common to all living organisms. It is thought that the rate of aging is influenced by diverse factors, in many cases related to the control of energy metabolism, i.e., the so-called pro-longevity effects of starvation. Translation, regarded as the main energy consumption process, lies at the center of interest, as it has a significant impact on the longevity phenomenon. It has been shown that perturbations in the translational apparatus may lead to a lower rate of aging. Therefore, the main aim of this study was to investigate aging in relation to the protein biosynthesis circuit, taking into account the uL11 ribosomal protein as a vital ribosomal element. To this end, we used set of yeast mutants with deleted single uL11A or uL11B genes and a double disruptant uL11AB mutant. We applied an integrated approach analyzing a broad range of biological parameters of yeast mutant cells, especially the longevity phenomenon, supplemented with biochemical and high throughput transcriptomic and metobolomic approaches. The analysis showed that the longevity phenomenon is not fully related to the commonly considered energy restriction effect, thus the slow-down of translation does not represent the sole source of aging. Additionally, we showed that uL11 can be classified as a moonlighting protein with extra-ribosomal function having cell-cycle regulatory potential.

ACS Style

Mateusz Mołoń; Eliza Molestak; Monika Kula-Maximenko; Przemysław Grela; Marek Tchórzewski. Ribosomal Protein uL11 as a Regulator of Metabolic Circuits Related to Aging and Cell Cycle. Cells 2020, 9, 1745 .

AMA Style

Mateusz Mołoń, Eliza Molestak, Monika Kula-Maximenko, Przemysław Grela, Marek Tchórzewski. Ribosomal Protein uL11 as a Regulator of Metabolic Circuits Related to Aging and Cell Cycle. Cells. 2020; 9 (7):1745.

Chicago/Turabian Style

Mateusz Mołoń; Eliza Molestak; Monika Kula-Maximenko; Przemysław Grela; Marek Tchórzewski. 2020. "Ribosomal Protein uL11 as a Regulator of Metabolic Circuits Related to Aging and Cell Cycle." Cells 9, no. 7: 1745.

Journal article
Published: 13 July 2019 in Cells
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The genome-wide duplication event observed in eukaryotes represents an interesting biological phenomenon, extending the biological capacity of the genome at the expense of the same genetic material. For example, most ribosomal proteins in Saccharomyces cerevisiae are encoded by a pair of paralogous genes. It is thought that gene duplication may contribute to heterogeneity of the translational machinery; however, the exact biological function of this event has not been clarified. In this study, we have investigated the functional impact of one of the duplicated ribosomal proteins, uL6, on the translational apparatus together with its consequences for aging of yeast cells. Our data show that uL6 is not required for cell survival, although lack of this protein decreases the rate of growth and inhibits budding. The uL6 protein is critical for the efficient assembly of the ribosome 60S subunit, and the two uL6 isoforms most likely serve the same function, playing an important role in the adaptation of translational machinery performance to the metabolic needs of the cell. The deletion of a single uL6 gene significantly extends the lifespan but only in cells with a high metabolic rate. We conclude that the maintenance of two copies of the uL6 gene enables the cell to cope with the high demands for effective ribosome synthesis.

ACS Style

Lidia Borkiewicz; Mateusz Mołoń; Eliza Molestak; Przemysław Grela; Patrycja Horbowicz-Drożdżal; Leszek Wawiórka; Marek Tchórzewski. Functional Analysis of the Ribosomal uL6 Protein of Saccharomyces cerevisiae. Cells 2019, 8, 718 .

AMA Style

Lidia Borkiewicz, Mateusz Mołoń, Eliza Molestak, Przemysław Grela, Patrycja Horbowicz-Drożdżal, Leszek Wawiórka, Marek Tchórzewski. Functional Analysis of the Ribosomal uL6 Protein of Saccharomyces cerevisiae. Cells. 2019; 8 (7):718.

Chicago/Turabian Style

Lidia Borkiewicz; Mateusz Mołoń; Eliza Molestak; Przemysław Grela; Patrycja Horbowicz-Drożdżal; Leszek Wawiórka; Marek Tchórzewski. 2019. "Functional Analysis of the Ribosomal uL6 Protein of Saccharomyces cerevisiae." Cells 8, no. 7: 718.

Review
Published: 27 April 2019 in Toxins
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Ricin belongs to the group of ribosome-inactivating proteins (RIPs), i.e., toxins that have evolved to provide particular species with an advantage over other competitors in nature. Ricin possesses RNA N-glycosidase activity enabling the toxin to eliminate a single adenine base from the sarcin-ricin RNA loop (SRL), which is a highly conserved structure present on the large ribosomal subunit in all species from the three domains of life. The SRL belongs to the GTPase associated center (GAC), i.e., a ribosomal element involved in conferring unidirectional trajectory for the translational apparatus at the expense of GTP hydrolysis by translational GTPases (trGTPases). The SRL represents a critical element in the GAC, being the main triggering factor of GTP hydrolysis by trGTPases. Enzymatic removal of a single adenine base at the tip of SRL by ricin blocks GTP hydrolysis and, at the same time, impedes functioning of the translational machinery. Here, we discuss the consequences of SRL depurination by ricin for ribosomal performance, with emphasis on the mechanistic model overview of the SRL modus operandi.

ACS Style

Przemysław Grela; Monika Szajwaj; Patrycja Horbowicz-Drożdżal; Marek Tchórzewski. How Ricin Damages the Ribosome. Toxins 2019, 11, 241 .

AMA Style

Przemysław Grela, Monika Szajwaj, Patrycja Horbowicz-Drożdżal, Marek Tchórzewski. How Ricin Damages the Ribosome. Toxins. 2019; 11 (5):241.

Chicago/Turabian Style

Przemysław Grela; Monika Szajwaj; Patrycja Horbowicz-Drożdżal; Marek Tchórzewski. 2019. "How Ricin Damages the Ribosome." Toxins 11, no. 5: 241.

Research article
Published: 20 September 2017 in PLOS ONE
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Frataxin is a highly conserved protein found in both prokaryotes and eukaryotes. It is involved in several central functions in cells, which include iron delivery to biochemical processes, such as heme synthesis, assembly of iron-sulfur clusters (ISC), storage of surplus iron in conditions of iron overload, and repair of ISC in aconitase. Frataxin from different organisms has been shown to undergo iron-dependent oligomerization. At least two different classes of oligomers, with different modes of oligomer packing and stabilization, have been identified. Here, we continue our efforts to explore the factors that control the oligomerization of frataxin from different organisms, and focus on E. coli frataxin CyaY. Using small-angle X-ray scattering (SAXS), we show that higher iron-to-protein ratios lead to larger oligomeric species, and that oligomerization proceeds in a linear fashion as a results of iron oxidation. Native mass spectrometry and online size-exclusion chromatography combined with SAXS show that a dimer is the most common form of CyaY in the presence of iron at atmospheric conditions. Modeling of the dimer using the SAXS data confirms the earlier proposed head-to-tail packing arrangement of monomers. This packing mode brings several conserved acidic residues into close proximity to each other, creating an environment for metal ion binding and possibly even mineralization. Together with negative-stain electron microscopy, the experiments also show that trimers, tetramers, pentamers, and presumably higher-order oligomers may exist in solution. Nano-differential scanning fluorimetry shows that the oligomers have limited stability and may easily dissociate at elevated temperatures. The factors affecting the possible oligomerization mode are discussed

ACS Style

Mostafa Fekry; Wessen Alshokry; Przemysław Grela; Marek Tchórzewski; Eva-Christina Ahlgren; Christopher A. Söderberg; Oleksandr Gakh; Grazia Isaya; Salam Al-Karadaghi. SAXS and stability studies of iron-induced oligomers of bacterial frataxin CyaY. PLOS ONE 2017, 12, e0184961 .

AMA Style

Mostafa Fekry, Wessen Alshokry, Przemysław Grela, Marek Tchórzewski, Eva-Christina Ahlgren, Christopher A. Söderberg, Oleksandr Gakh, Grazia Isaya, Salam Al-Karadaghi. SAXS and stability studies of iron-induced oligomers of bacterial frataxin CyaY. PLOS ONE. 2017; 12 (9):e0184961.

Chicago/Turabian Style

Mostafa Fekry; Wessen Alshokry; Przemysław Grela; Marek Tchórzewski; Eva-Christina Ahlgren; Christopher A. Söderberg; Oleksandr Gakh; Grazia Isaya; Salam Al-Karadaghi. 2017. "SAXS and stability studies of iron-induced oligomers of bacterial frataxin CyaY." PLOS ONE 12, no. 9: e0184961.

Journal article
Published: 01 September 2017 in Molecular and Cellular Biology
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The P-stalk represents a vital element within the ribosomal GTPase-associated center, which represents a landing platform for translational GTPases. The eukaryotic P-stalk exists as a uL10-(P1-P2) 2 pentameric complex, which contains five identical C-terminal domains, one within each protein, and the presence of only one such element is sufficient to stimulate factor-dependent GTP hydrolysis in vitro and to sustain cell viability. The functional contribution of the P-stalk to the performance of the translational machinery in vivo , especially the role of P-protein multiplication, has never been explored. Here, we show that ribosomes depleted of P1/P2 proteins exhibit reduced translation fidelity at elongation and termination steps. The elevated rate of the decoding error is inversely correlated with the number of the P-proteins present on the ribosome. Unexpectedly, the lack of P1/P2 has little effect in vivo on the efficiency of other translational GTPase (trGTPase)-dependent steps of protein synthesis, including translocation. We have shown that loss of accuracy of decoding caused by P1/P2 depletion is the major cause of translation slowdown, which in turn affects the metabolic fitness of the yeast cell. We postulate that the multiplication of P-proteins is functionally coupled with the qualitative aspect of ribosome action, i.e., the recoding phenomenon shaping the cellular proteome.

ACS Style

Leszek Wawiórka; Eliza Molestak; Monika Szajwaj; Barbara Michalec-Wawiórka; Mateusz Mołoń; Lidia Borkiewicz; Przemysław Grela; Aleksandra Boguszewska; Marek Tchórzewski. Multiplication of Ribosomal P-Stalk Proteins Contributes to the Fidelity of Translation. Molecular and Cellular Biology 2017, 37, e00060-17 .

AMA Style

Leszek Wawiórka, Eliza Molestak, Monika Szajwaj, Barbara Michalec-Wawiórka, Mateusz Mołoń, Lidia Borkiewicz, Przemysław Grela, Aleksandra Boguszewska, Marek Tchórzewski. Multiplication of Ribosomal P-Stalk Proteins Contributes to the Fidelity of Translation. Molecular and Cellular Biology. 2017; 37 (17):e00060-17.

Chicago/Turabian Style

Leszek Wawiórka; Eliza Molestak; Monika Szajwaj; Barbara Michalec-Wawiórka; Mateusz Mołoń; Lidia Borkiewicz; Przemysław Grela; Aleksandra Boguszewska; Marek Tchórzewski. 2017. "Multiplication of Ribosomal P-Stalk Proteins Contributes to the Fidelity of Translation." Molecular and Cellular Biology 37, no. 17: e00060-17.

Journal article
Published: 17 July 2017 in Scientific Reports
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The eukaryotic P-stalk contains two P1-P2 protein dimers with a conserved C- terminal domain (CTD) critical for the interaction with external factors. To understand the role of the individual CTD of human P1/P2 proteins, we examined the interaction of reconstituted human P-protein complexes and C-terminally truncated forms with ricin A chain (RTA), which binds to the stalk to depurinate the sarcin/ricin loop (SRL). The interaction between P-protein complexes and RTA was examined by surface plasmon resonance, isothermal titration calorimetry, microscale thermophoresis and bio-layer interferometry. The P1-P2 heterodimer missing a CTD on P2 was able to bind RTA. In contrast, the P1-P2 heterodimer missing the CTD of P1 protein displayed almost no binding toward RTA. Very low interaction was detected between RTA and the non-truncated P2-P2 homodimer, suggesting that the structural architecture of the P1-P2 heterodimer is critical for binding RTA. The reconstituted pentameric human stalk complex had higher affinity for RTA than the P1-P2 dimer. Deletion of P1 CTD, but not P2 CTD reduced the affinity of the pentamer for RTA. These results highlight the importance of the heterodimeric organization of P1-P2 in the human stalk pentamer and functional non-equivalence of the individual P-protein CTDs in the interaction with RTA.

ACS Style

Przemysław Grela; Xiao-Ping Li; Patrycja Horbowicz; Monika Dźwierzyńska; Marek Tchórzewski; Nilgun E. Tumer. Human ribosomal P1-P2 heterodimer represents an optimal docking site for ricin A chain with a prominent role for P1 C-terminus. Scientific Reports 2017, 7, 5608 .

AMA Style

Przemysław Grela, Xiao-Ping Li, Patrycja Horbowicz, Monika Dźwierzyńska, Marek Tchórzewski, Nilgun E. Tumer. Human ribosomal P1-P2 heterodimer represents an optimal docking site for ricin A chain with a prominent role for P1 C-terminus. Scientific Reports. 2017; 7 (1):5608.

Chicago/Turabian Style

Przemysław Grela; Xiao-Ping Li; Patrycja Horbowicz; Monika Dźwierzyńska; Marek Tchórzewski; Nilgun E. Tumer. 2017. "Human ribosomal P1-P2 heterodimer represents an optimal docking site for ricin A chain with a prominent role for P1 C-terminus." Scientific Reports 7, no. 1: 5608.

Journal article
Published: 01 May 2017 in Ornithological Applications
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Knowledge about interactions between habitat quality and individual condition is vital for understanding ecological processes. We examined variation in commonly used hematological indices and body mass in incubating and chick-rearing Red-necked Grebes (Podiceps grisegena) in Poland breeding under contrasting foraging conditions in ponds used for aquaculture. Grebes used either ponds with low prey availability in the early breeding period and abundant food in the brood-rearing stage (high-quality habitat), or ponds stocked with fish attractive as prey to spring-arriving adults yet later unavailable to chicks due to size constraints (low-quality habitat acting as an ecological trap). Parental effort, measured as the proportion of time allocated to food acquisition and provisioning of chicks, was greater in the low-quality habitat, although adult grebes spent less time foraging there than in the high-quality habitat early in the season. Scaled body mass was higher in incubating grebes than in brood-rearing grebes, regardless of habitat quality. Breeding stage by habitat interactions were significant for hemoglobin, erythrocyte count (RBC), mean corpuscular volume (MCV), and mean corpuscular hemoglobin (MCH), with greater MCV and MCH values (by 20–34%, on average) during brood-rearing than during incubation in the low-quality habitat, and the opposite pattern (by 15–16%) in the high-quality habitat. These hematological responses signify changes in physiological stress associated with parental workloads. Higher MCV and MCH levels (by 22–31%) during the chick-rearing stage in the low-quality compared with the high-quality habitat indicate that parents incurred physiological costs of breeding under ecological trap conditions. No significant effects were found on leucocyte indices (total leucocyte count and heterophil/lymphocyte ratio). We consider the hemoglobin- and RBC-related parameters to be promising indicators of habitat-driven stress associated with prolonged physical exertion, especially when multiple metrics are employed. RESUMEN El conocimiento sobre las interacciones entre la calidad del hábitat y la condición de los individuos es vital para entender los procesos ecológicos. Examinamos la variación en los índices hematológicos comúnmente usados y en la masa corporal de individuos reproductivos de Podiceps grisegena incubando y criando polluelos bajo condiciones de forrajeo contrastantes en estanques de acuicultura en Polonia. P. grisegena utilizó estanques con baja disponibilidad de presas a inicios del período reproductivo y con alta abundancia de alimento en la etapa de cría (hábitats de alta calidad) o estanques llenos de peces atractivos como presas para los adultos que llegan en primavera aunque luego no disponibles para los polluelos debido a restricciones de tamaño (hábitat de baja calidad que actúa como una trampa ecológica). El esfuerzo parental, medido como la proporción de tiempo asignado a la adquisición de alimento y al aprovisionamiento de los polluelos, fue mayor en el hábitat de baja calidad, aunque a inicios de la temporada los adultos pasaron menos tiempo forrajeando en este hábitat que en el hábitat de alta calidad. La masa corporal llevada a escala fue más alta en los individuos de P. grisegena que estaban incubando, que en los que estaban criando, independientemente del tipo de hábitat. La etapa reproductiva por las interacciones del hábitat fueron significativas para la hemoglobina, los conteos de eritrocitos (CE), el volumen corpuscular promedio (VCP) y la hemoglobina corpuscular promedio (HCP), con valores más altos de VCP y HCP (en promedio por 20–34%) durante la cría que durante la incubación en el hábitat de baja calidad y un patrón opuesto (por 15–16%) en el hábitat de alta calidad. Estas respuestas hematológicas significan cambios en el estrés fisiológico asociado con las cargas de trabajo de los padres. Mayores niveles de VCP y HCP (por 22–31%) durante la etapa de polluelo en el hábitat de baja calidad indicaron que los padres tuvieron costos fisiológicos derivados de la reproducción en las condiciones de trampa ecológica. No se encontraron efectos significativos en los índices de leucocitos (conteo total de leucocitos y cociente heterófilo/linfocito). Consideramos que los parámetros relacionados con la hemoglobina y el CE son indicadores promisorios del estrés causado por el hábitat asociado con un esfuerzo físico prolongado, especialmente cuando se emplean múltiples métricas. Palabras clave: costos reproductivos, ecofisiología, esfuerzo de forrajeo, índices de condición, selección de hábitat

ACS Style

Janusz Kloskowski; Elżbieta Kaczanowska; Jarosław Krogulec; Przemysław Grela. Hematological indicators of habitat quality: Erythrocyte parameters reflect greater parental effort of Red-necked Grebes under ecological trap conditions. Ornithological Applications 2017, 119, 239 -250.

AMA Style

Janusz Kloskowski, Elżbieta Kaczanowska, Jarosław Krogulec, Przemysław Grela. Hematological indicators of habitat quality: Erythrocyte parameters reflect greater parental effort of Red-necked Grebes under ecological trap conditions. Ornithological Applications. 2017; 119 (2):239-250.

Chicago/Turabian Style

Janusz Kloskowski; Elżbieta Kaczanowska; Jarosław Krogulec; Przemysław Grela. 2017. "Hematological indicators of habitat quality: Erythrocyte parameters reflect greater parental effort of Red-necked Grebes under ecological trap conditions." Ornithological Applications 119, no. 2: 239-250.

Comparative study
Published: 25 April 2014 in Biochemical Journal
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Wild-type yeast ribosomes and ribosomes bearing only the P1B–P2A dimer bound to the ricin A chain better and were more susceptible to depurination and toxicity than ribosomes bearing only the P1A–P2B dimer, indicating that the two stalk dimers differ in their function.

ACS Style

Przemysław Grela; Xiao-Ping Li; Marek Tchórzewski; Nilgun E. Tumer. Functional divergence between the two P1–P2 stalk dimers on the ribosome in their interaction with ricin A chain. Biochemical Journal 2014, 460, 59 -69.

AMA Style

Przemysław Grela, Xiao-Ping Li, Marek Tchórzewski, Nilgun E. Tumer. Functional divergence between the two P1–P2 stalk dimers on the ribosome in their interaction with ricin A chain. Biochemical Journal. 2014; 460 (1):59-69.

Chicago/Turabian Style

Przemysław Grela; Xiao-Ping Li; Marek Tchórzewski; Nilgun E. Tumer. 2014. "Functional divergence between the two P1–P2 stalk dimers on the ribosome in their interaction with ricin A chain." Biochemical Journal 460, no. 1: 59-69.

Journal article
Published: 01 October 2013 in Journal of Biological Chemistry
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Ricin inhibits protein synthesis by depurinating the α-sarcin/ricin loop (SRL). Ricin holotoxin does not inhibit translation unless the disulfide bond between the A (RTA) and B (RTB) subunits is reduced. Ricin holotoxin did not bind ribosomes or depurinate them but could depurinate free RNA. When RTA is separated from RTB, arginine residues located at the interface are exposed to the solvent. Because this positively charged region, but not the active site, is blocked by RTB, we mutated arginine residues at or near the interface of RTB to determine if they are critical for ribosome binding. These variants were structurally similar to wild type RTA but could not bind ribosomes. Their Km values and catalytic rates (kcat) for an SRL mimic RNA were similar to those of wild type, indicating that their activity was not altered. However, they showed an up to 5-fold increase in Km and up to 38-fold decrease in kcat toward ribosomes. These results suggest that the stalk binding stimulates the catalysis of ribosome depurination by RTA. The mutated arginines have side chains behind the active site cleft, indicating that the ribosome binding surface of RTA is on the opposite side of the surface that interacts with the SRL. We propose that stalk binding stimulates the catalysis of ribosome depurination by orienting the active site of RTA toward the SRL and thereby allows docking of the target adenine into the active site. This model may apply to the translation factors that interact with the stalk.

ACS Style

Xiao-Ping Li; Peter C. Kahn; Jennifer Nielsen Kahn; Przemysław Grela; Nilgun E. Tumer. Arginine Residues on the Opposite Side of the Active Site Stimulate the Catalysis of Ribosome Depurination by Ricin A Chain by Interacting with the P-protein Stalk. Journal of Biological Chemistry 2013, 288, 30270 -30284.

AMA Style

Xiao-Ping Li, Peter C. Kahn, Jennifer Nielsen Kahn, Przemysław Grela, Nilgun E. Tumer. Arginine Residues on the Opposite Side of the Active Site Stimulate the Catalysis of Ribosome Depurination by Ricin A Chain by Interacting with the P-protein Stalk. Journal of Biological Chemistry. 2013; 288 (42):30270-30284.

Chicago/Turabian Style

Xiao-Ping Li; Peter C. Kahn; Jennifer Nielsen Kahn; Przemysław Grela; Nilgun E. Tumer. 2013. "Arginine Residues on the Opposite Side of the Active Site Stimulate the Catalysis of Ribosome Depurination by Ricin A Chain by Interacting with the P-protein Stalk." Journal of Biological Chemistry 288, no. 42: 30270-30284.

Original article
Published: 22 August 2012 in The FEBS Journal
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Ricin A‐chain (RTA) depurinates the sarcin–ricin loop of 28S ribosomal RNA and inhibits protein synthesis in mammalian cells. In yeast, the ribosomal stalk facilitates the interaction of RTA with the ribosome and subsequent depurination. Despite homology between the stalk structures from yeast and humans, there are notable differences. The human ribosomal stalk contains two identical heterodimers of P1 and P2 bound to P0, whereas the yeast stalk consists of two different heterodimers, P1α–P2β and P2α–P1β, bound to P0. RTA exhibits higher activity towards mammalian ribosomes than towards ribosomes from other organisms, suggesting that the mode of interaction with ribosomes may vary. Here, we examined whether the human ribosomal stalk proteins facilitate the interaction of RTA with human ribosomes and subsequent depurination of the sarcin–ricin loop. Using small interfering RNA‐mediated knockdown of P1/P2 expression in human cells, we demonstrated that the depurination activity of RTA is lower when P1 and P2 levels are reduced. Biacore analysis showed that ribosomes from P1/P2‐depleted cells have a reduced ability to bind RTA, which correlates with reduced depurination activity both in vitro and inside cells. RTA interacts directly with recombinant human P1–P2 dimer, further demonstrating the importance of human P1 and P2 in enabling RTA to bind and depurinate human ribosomes. Structured digital abstract P2, P1 and RTA physically interact by surface plasmon resonance (View interaction) P2, P1 and RTA physically interact by anti bait coimmunoprecipitation (View interaction)

ACS Style

Kerrie L. May; Xiao-Ping Li; Francisco Martínez-Azorín; Juan P. G. Ballesta; Przemysław Grela; Marek Tchórzewski; Nilgun E. Tumer. The P1/P2 proteins of the human ribosomal stalk are required for ribosome binding and depurination by ricin in human cells. The FEBS Journal 2012, 279, 3925 -3936.

AMA Style

Kerrie L. May, Xiao-Ping Li, Francisco Martínez-Azorín, Juan P. G. Ballesta, Przemysław Grela, Marek Tchórzewski, Nilgun E. Tumer. The P1/P2 proteins of the human ribosomal stalk are required for ribosome binding and depurination by ricin in human cells. The FEBS Journal. 2012; 279 (20):3925-3936.

Chicago/Turabian Style

Kerrie L. May; Xiao-Ping Li; Francisco Martínez-Azorín; Juan P. G. Ballesta; Przemysław Grela; Marek Tchórzewski; Nilgun E. Tumer. 2012. "The P1/P2 proteins of the human ribosomal stalk are required for ribosome binding and depurination by ricin in human cells." The FEBS Journal 279, no. 20: 3925-3936.

Comparative study
Published: 11 May 2012 in Biochemical Journal
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The ribosomal stalk of the 60S subunit has been shown to play a crucial role in all steps of protein synthesis, but its structure and exact molecular function remain an unanswered question. In the present study, we show the low-resolution models of the solution structure of the yeast ribosomal stalk, composed of five proteins, P0–(P1–P2)2. The model of the pentameric stalk complex determined by small-angle X-ray scattering reveals an elongated shape with a maximum length of 13 nm. The model displays three distinct lobes, which may correspond to the individual P1–P2 heterodimers anchored to the C-terminal domain of the P0 protein.

ACS Style

Przemysław Grela; Michal J. Gajda; Jean-Paul Armache; Roland Beckmann; Dawid Krokowski; Dmitri Svergun; Nikodem Grankowski; Marek Tchórzewski. Solution structure of the natively assembled yeast ribosomal stalk determined by small-angle X-ray scattering. Biochemical Journal 2012, 444, 205 -209.

AMA Style

Przemysław Grela, Michal J. Gajda, Jean-Paul Armache, Roland Beckmann, Dawid Krokowski, Dmitri Svergun, Nikodem Grankowski, Marek Tchórzewski. Solution structure of the natively assembled yeast ribosomal stalk determined by small-angle X-ray scattering. Biochemical Journal. 2012; 444 (2):205-209.

Chicago/Turabian Style

Przemysław Grela; Michal J. Gajda; Jean-Paul Armache; Roland Beckmann; Dawid Krokowski; Dmitri Svergun; Nikodem Grankowski; Marek Tchórzewski. 2012. "Solution structure of the natively assembled yeast ribosomal stalk determined by small-angle X-ray scattering." Biochemical Journal 444, no. 2: 205-209.

Research article
Published: 01 January 2012 in Behaviour
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In monogamous birds, early male parental effort, such as nest building, may serve as a post-mating sexually-selected display allowing female assessment of male quality. We examined the functional significance of male nest building and the potential role of nest size as a sexually-selected signal in the red-necked grebe ( Podiceps grisegena), a species with high mate fidelity. Time-activity budgets showed that no behaviour was performed exclusively by one sex in the pre-laying period, but males spent significantly more time nest building and were more often involved in aggressive intra- and interspecific interactions. Nest building in pairs attempting a second brood was also performed predominantly by males. Greater participation in nest construction by males allowed females to allocate more time to self-maintenance activities in the period prior to egg-laying. The positive relationship found between the relative contribution of males to nest building and later to brood provisioning indicates that male nest building is an honest indicator of future paternal effort. Males obtained copulations solicited by females proportionally to the time spent on nest building, and the extent of male participation in nest construction was of importance for explaining variation in clutch size. Nest size itself is not very likely to be sexually selected in red-necked grebes, as it was found to depend on nest site conditions such as water depth and exposure to wave action. We suggest that greater investment of males in energetically demanding pre-laying activities is functionally similar to post-mating courtship feeding; it constitutes males’ indirect contribution to clutch production and may help to negotiate the relative investment each sex makes in the different stages of the breeding cycle. The results support the idea that, in monogamous birds, naturally selected male characters related to parental care may evolve into important sexual signals to females, although not into extreme displays.

ACS Style

Michał Gąska; Przemysław Grela; Janusz Kloskowski. The role of male nest building in post-mating sexual selection in the monogamous red-necked grebe. Behaviour 2012, 149, 81 -98.

AMA Style

Michał Gąska, Przemysław Grela, Janusz Kloskowski. The role of male nest building in post-mating sexual selection in the monogamous red-necked grebe. Behaviour. 2012; 149 (1):81-98.

Chicago/Turabian Style

Michał Gąska; Przemysław Grela; Janusz Kloskowski. 2012. "The role of male nest building in post-mating sexual selection in the monogamous red-necked grebe." Behaviour 149, no. 1: 81-98.

Journal article
Published: 01 December 2010 in Journal of Biological Chemistry
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Ribosome inactivating proteins (RIPs) depurinate a universally conserved adenine in the α-sarcin/ricin loop (SRL) and inhibit protein synthesis at the translation elongation step. We previously showed that ribosomal stalk is required for depurination of the SRL by ricin toxin A chain (RTA). The interaction between RTA and ribosomes was characterized by a two-step binding model, where the stalk structure could be considered as an important interacting element. Here, using purified yeast ribosomal stalk complexes assembled in vivo, we show a direct interaction between RTA and the isolated stalk complex. Detailed kinetic analysis of these interactions in real time using surface plasmon resonance (SPR) indicated that there is only one type of interaction between RTA and the ribosomal stalk, which represents one of the two binding steps of the interaction with ribosomes. Interactions of RTA with the isolated stalk were relatively insensitive to salt, indicating that nonelectrostatic interactions were dominant. We compared the interaction of RTA with the full pentameric stalk complex containing two pairs of P1/P2 proteins with its interaction with the trimeric stalk complexes containing only one pair of P1/P2 and found that the rate of association of RTA with the pentamer was higher than with either trimer. These results demonstrate that the stalk is the main landing platform for RTA on the ribosome and that pentameric organization of the stalk accelerates recruitment of RTA to the ribosome for depurination. Our results suggest that multiple copies of the stalk proteins might also increase the scavenging ability of the ribosome for the translational GTPases.

ACS Style

Xiao-Ping Li; Przemyslaw Grela; Dawid Krokowski; Marek Tchórzewski; Nilgun E. Tumer. Pentameric Organization of the Ribosomal Stalk Accelerates Recruitment of Ricin A Chain to the Ribosome for Depurination. Journal of Biological Chemistry 2010, 285, 41463 -41471.

AMA Style

Xiao-Ping Li, Przemyslaw Grela, Dawid Krokowski, Marek Tchórzewski, Nilgun E. Tumer. Pentameric Organization of the Ribosomal Stalk Accelerates Recruitment of Ricin A Chain to the Ribosome for Depurination. Journal of Biological Chemistry. 2010; 285 (53):41463-41471.

Chicago/Turabian Style

Xiao-Ping Li; Przemyslaw Grela; Dawid Krokowski; Marek Tchórzewski; Nilgun E. Tumer. 2010. "Pentameric Organization of the Ribosomal Stalk Accelerates Recruitment of Ricin A Chain to the Ribosome for Depurination." Journal of Biological Chemistry 285, no. 53: 41463-41471.

Comparative study
Published: 31 May 2010 in The International Journal of Biochemistry & Cell Biology
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The Mrt4 protein, showing extensive sequence similarity to the ribosomal P0 protein, is classified as a ribosomal P0-like protein and acts as a trans-acting factor which modulates the assembly of the pre-60S particle. In this report we investigated the biological nature of the human Mrt4 protein. First, we constructed a series of hybrid hMrt4-P0 proteins by replacing various domains of the P0 protein with corresponding protein fragments from hMrt4. We found that hMrt4 binds to the same site on the large ribosomal subunit as does P0, but despite the sequence homology it is not able to functionally complement the lack of P0. Using fluorescence microscopy and biochemical approaches we also show that hMrt4 occupies predominantly the nucleolar compartment, in contrast to P0 and P1/P2, which are located in the cytoplasm. The nucleolar accumulation of hMrt4 does not depend on a specific nucleolus localization signal, but rather occurs via interaction with established nucleolar components such as rRNA; however, nuclear import of hMrt4 is dependent on a short sequence in the N-terminal part of the protein. Functional analysis with specific inhibitors, actinomycin D and leptomycin B, showed that hMrt4 is a trans-acting factor involved in ribosome maturation, with nucleus-cytoplasm shuttling capacity.

ACS Style

Barbara Michalec; Dawid Krokowski; Przemysław Grela; Leszek Wawiórka; Justyna Sawa-Makarska; Nikodem Grankowski; Marek Tchórzewski. Subcellular localization of ribosomal P0-like protein MRT4 is determined by its N-terminal domain. The International Journal of Biochemistry & Cell Biology 2010, 42, 736 -748.

AMA Style

Barbara Michalec, Dawid Krokowski, Przemysław Grela, Leszek Wawiórka, Justyna Sawa-Makarska, Nikodem Grankowski, Marek Tchórzewski. Subcellular localization of ribosomal P0-like protein MRT4 is determined by its N-terminal domain. The International Journal of Biochemistry & Cell Biology. 2010; 42 (5):736-748.

Chicago/Turabian Style

Barbara Michalec; Dawid Krokowski; Przemysław Grela; Leszek Wawiórka; Justyna Sawa-Makarska; Nikodem Grankowski; Marek Tchórzewski. 2010. "Subcellular localization of ribosomal P0-like protein MRT4 is determined by its N-terminal domain." The International Journal of Biochemistry & Cell Biology 42, no. 5: 736-748.

Journal article
Published: 19 May 2010 in Biophysical Journal
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Ribosomal protein L12 is a two-domain protein that forms dimers mediated by its N-terminal domains. A 20-residue linker separates the N- and C-terminal domains. This linker results in a three-lobe topology with significant flexibility, known to be critical for efficient translation. Here we present an ensemble model of spatial distributions and correlation times for the domain reorientations of L12 that reconciles experimental data from small-angle x-ray scattering and nuclear magnetic resonance. We generated an ensemble of L12 conformations in which the structure of each domain is fixed but the domain orientations are variable. The ensemble reproduces the small-angle x-ray scattering data and the optimized correlation times of its reorientational eigenmodes fit the 15N relaxation data. The ensemble model reveals intrinsic conformational properties of L12 that help explain its function on the ribosome. The two C-terminal domains sample a large volume and extend further away from the ribosome anchor than expected for a random-chain linker, indicating that the flexible linker has residual order. Furthermore, the distances between each C-terminal domain and the anchor are anticorrelated, indicating that one of them is more retracted on average. We speculate that these properties promote the function of L12 to recruit translation factors and control their activity on the ribosome.

ACS Style

Pau Bernadó; Kristofer Modig; Przemysław Grela; Dmitri I. Svergun; Marek Tchorzewski; Miquel Pons; Mikael Akke. Structure and Dynamics of Ribosomal Protein L12: An Ensemble Model Based on SAXS and NMR Relaxation. Biophysical Journal 2010, 98, 2374 -2382.

AMA Style

Pau Bernadó, Kristofer Modig, Przemysław Grela, Dmitri I. Svergun, Marek Tchorzewski, Miquel Pons, Mikael Akke. Structure and Dynamics of Ribosomal Protein L12: An Ensemble Model Based on SAXS and NMR Relaxation. Biophysical Journal. 2010; 98 (10):2374-2382.

Chicago/Turabian Style

Pau Bernadó; Kristofer Modig; Przemysław Grela; Dmitri I. Svergun; Marek Tchorzewski; Miquel Pons; Mikael Akke. 2010. "Structure and Dynamics of Ribosomal Protein L12: An Ensemble Model Based on SAXS and NMR Relaxation." Biophysical Journal 98, no. 10: 2374-2382.

Research article
Published: 08 January 2010 in Biochemistry
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The landing platform for the translational GTPases is located on the 60S ribosomal subunit and is referred to as a GTPase-associated center. The most distinctive feature of this center is an oligomeric complex, the stalk, responsible for the recruitment of translation factors and stimulation of translation factor-dependent GTP hydrolysis. In eukaryotes, the stalk has been investigated in vitro and in vivo, but most information available concerns its individual components only. In the present study, we provide an insight into the biophysical nature of the native stalk isolated from the yeast Saccharomyces cerevisiae. Using fluorescence, circular dichroism, and mass spectrometry analyses, we were able to characterize the natively formed yeast stalk, casting new light on the oligomeric properties of the complex and its quaternary topology, showing that folding and assembly are coupled processes. The pentameric stalk is an exceptionally stable structure with the protein core composed of P0, P1A, and P2B proteins and less tightly bound P1B and P2A capable of dissociating from the stalk core. We obtained also the whole picture of the posttranslational modifications at the logarithmic phase of yeast growth, using mass spectrometry approach, where P proteins are phosphorylated at a single serine residue, P0 may accept two phosphate groups, and P1A none. Additionally, only P1B undergoes N-terminal acetylation after prior methionine removal.

ACS Style

Przemysław Grela; Dawid Krokowski; Yuliya Gordiyenko; Daniel Krowarsch; Carol Robinson; Jacek Otlewski; Nikodem Grankowski; Marek Tchórzewski. Biophysical Properties of the Eukaryotic Ribosomal Stalk. Biochemistry 2010, 49, 924 -933.

AMA Style

Przemysław Grela, Dawid Krokowski, Yuliya Gordiyenko, Daniel Krowarsch, Carol Robinson, Jacek Otlewski, Nikodem Grankowski, Marek Tchórzewski. Biophysical Properties of the Eukaryotic Ribosomal Stalk. Biochemistry. 2010; 49 (5):924-933.

Chicago/Turabian Style

Przemysław Grela; Dawid Krokowski; Yuliya Gordiyenko; Daniel Krowarsch; Carol Robinson; Jacek Otlewski; Nikodem Grankowski; Marek Tchórzewski. 2010. "Biophysical Properties of the Eukaryotic Ribosomal Stalk." Biochemistry 49, no. 5: 924-933.

Article
Published: 09 July 2008 in Journal of Molecular Evolution
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The GTPase center of the large ribosomal subunit, being a landing platform for translation factors, and regarded as one of the oldest structures in the ribosome, is a universally conserved structure in all domains of life. It is thought that this structure could be responsible for the major breakthrough on the way to the RNA/protein world, because its appearance would have dramatically increased the rate and accuracy of protein synthesis. The major part of this center is recognized as a distinct structural entity, called the stalk. The main functional part of the stalk in all domains of life is composed of small L12/P proteins, which are believed to form an evolutionarily conserved group. However, some data indicate that the bacterial and archaeo/eukaryal proteins are not related to each other structurally, and only a functional relationship may be clearly recognized. To clarify this point, we performed a comprehensive comparative analysis of the L12/P proteins from the three domains of life. The results show that bacterial and archaeo/eukaryal L12/P-proteins are not structurally related and, therefore, might not be linked evolutionarily either. Consequently, these proteins should be regarded as analogous rather than homologous systems and probably appeared on the ribosomal particle in two independent events in the course of evolution.

ACS Style

Przemysław Grela; Pau Bernado; Dmitri Svergun; Jan Kwiatowski; Dariusz Abramczyk; Nikodem Grankowski; Marek Tchórzewski. Structural Relationships Among the Ribosomal Stalk Proteins from the Three Domains of Life. Journal of Molecular Evolution 2008, 67, 154 -167.

AMA Style

Przemysław Grela, Pau Bernado, Dmitri Svergun, Jan Kwiatowski, Dariusz Abramczyk, Nikodem Grankowski, Marek Tchórzewski. Structural Relationships Among the Ribosomal Stalk Proteins from the Three Domains of Life. Journal of Molecular Evolution. 2008; 67 (2):154-167.

Chicago/Turabian Style

Przemysław Grela; Pau Bernado; Dmitri Svergun; Jan Kwiatowski; Dariusz Abramczyk; Nikodem Grankowski; Marek Tchórzewski. 2008. "Structural Relationships Among the Ribosomal Stalk Proteins from the Three Domains of Life." Journal of Molecular Evolution 67, no. 2: 154-167.

Journal article
Published: 04 November 2007 in Journal of Biochemistry
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ACS Style

Przemysław Grela; Justyna Sawa-Makarska; Yuliya Gordiyenko; Carol V. Robinson; Nikodem Grankowski; Marek Tchórzewski. Structural Properties of the Human Acidic Ribosomal P Proteins Forming the P1-P2 Heterocomplex. Journal of Biochemistry 2007, 143, 169 -177.

AMA Style

Przemysław Grela, Justyna Sawa-Makarska, Yuliya Gordiyenko, Carol V. Robinson, Nikodem Grankowski, Marek Tchórzewski. Structural Properties of the Human Acidic Ribosomal P Proteins Forming the P1-P2 Heterocomplex. Journal of Biochemistry. 2007; 143 (2):169-177.

Chicago/Turabian Style

Przemysław Grela; Justyna Sawa-Makarska; Yuliya Gordiyenko; Carol V. Robinson; Nikodem Grankowski; Marek Tchórzewski. 2007. "Structural Properties of the Human Acidic Ribosomal P Proteins Forming the P1-P2 Heterocomplex." Journal of Biochemistry 143, no. 2: 169-177.