Person: Martínez Del Pozo, Álvaro
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First Name
Álvaro
Last Name
Martínez Del Pozo
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Ciencias Químicas
Department
Bioquímica y Biología Molecular
Area
Bioquímica y Biología Molecular
Identifiers
13 results
Search Results
Now showing 1 - 10 of 13
Item Der p 1‑based immunotoxin as potential tool for the treatment of dust mite respiratory allergy(Scientific Reports, 2020) Lázaro‑Gorines, Rodrigo; López Rodríguez, Juan Carlos; Benedé Pérez, Sara; González, Miguel; Mayorga, Cristobalina; Vogel, Lothar; Martínez Del Pozo, Álvaro; Lacadena García-Gallo, Francisco Javier; Villalba Díaz, María TeresaImmunotoxins appear as promising therapeutic molecules, alternative to allergen-specifcimmunotherapy. In this work, we achieved the development of a protein chimera able to promote specifc cell death on efector cells involved in the allergic reaction. Der p 1 allergen was chosen as cell-targeting domain and the powerful ribotoxin α-sarcin as the toxic moiety. The resultant construction, named proDerp1αS, was produced and purifed from the yeast Pichia pastoris. Der p 1-protease activity and α-sarcin ribonucleolytic action were efectively conserved in proDerp1αS. Immunotoxin impact was assayed by using efector cells sensitized with house dust mite-allergic sera. Cell degranulation and death, triggered by proDerp1αS, was exclusively observed on Der p 1 sera sensitized-humRBL-2H3 cells, but not when treated with non-allergic sera. Most notably, equivalent IgE-binding and degranulation were observed with both proDerp1αS construct and native Der p 1 when using purifed basophils from sensitized patients. However, proDerp1αS did not cause any cytotoxic efect on these cells, apparently due to its lack of internalization after their surface IgEbinding, showing the complex in vivo panorama governing allergic reactions. In conclusion, herein we present proDerp1αS as a proof of concept for a potential and alternative new designs of therapeutic tools for allergies. Development of new, and more specifc, second-generation of immunotoxins following proDerp1αS, is further discussed- Project number: 117
Item I.amAble: aprendizaje e inclusión educativa mediante talleres científicos(2020) Herrero Domínguez, Santiago; Martínez Del Pozo, Álvaro; Mancheño Real, María José; Osío Barcina, José De Jesús; Nacenta Torres, Pablo; Bárcena Espelleta, Araceli; Rubio Lago, Luis; Álvarez Serrano, Inmaculada; Cortés Gil, Raquel; González Prieto, Rodrigo; Torrecilla Manresa, Sofia; Cilleros Prados, Olga; Sobrino Díaz, María Lourdes; Bartolomé Vílchez, Javier; Cortijo Montes, Miguel; Coloma Manjón-Cabeza, Isabel; Catalán Torrecilla, Cristina; Guerrero Martínez, Andrés; Martínez Madrid, Carmen Belén; Martínez Ruiz, María Paloma; Méndez Pozo, Gonzalo Rubén; Ranchal Sánchez, Rocío; Priego Bermejo, José Luis; Maestre Varea, David; Desvoyes, Bénedicte; Gutiérrez Franco, Yanna María; Arribas Fernández, Paula; Julián Cortés, ÁlvaroI.amAble ha ofrecido a estudiantes universitarios de física, química, veterinaria, biología y educación la oportunidad de complementar su formación mediante el diseño, la realización y la evaluación de talleres científicos que faciliten la inclusión de personas con diversidad cognitiva. Los talleres han sido diseñados por el alumnado universitario de ciencias y perfilados por estudiantes de educación para ser llevados a cabo por alumnado preuniversitario en parejas, de forma que un miembro pertenezca a un centro de secundaria ordinario y el otro miembro a un centro de educación especial. Aquellos talleres que se han considerado más adecuados por su adaptabilidad se han llevado a la práctica guiados por estudiantes de ciencias y de educación. Los miembros del proyecto, que incluyen representantes de todos los estamentos universitarios, han supervisado todas las tareas descritas anteriormente. Además de los miembros de la Universidad Complutense, también figuran personas voluntarias de otras instituciones científicas y educativas. El alumnado universitario ha tenido la posibilidad, no sólo de asentar y profundizar algunos contenidos científicos o poner en práctica algunas de las enseñanzas adquiridas, sino también de desarrollar su empatía, su capacidad de comunicar e improvisar y de adaptarse a un público heterogéneo. Ello ha mejorado sus perspectivas laborales, especialmente dentro de la educación formal e informal (animación sociocultural, museos científicos...). Además, han contribuido a facilitar la inclusión educativa de las personas con diversidad funcional y a mejorar la cultura científica de la sociedad. Con este proyecto, inspirado en la metodología Aprendizaje-Servicio (ApS), se ha pretendido también mejorar la accesibilidad a las experiencias y contenidos científicos y facilitar la inclusión educativa de las personas con diversidad funcional, especialmente diversidad cognitiva o intelectual. En la primera edición de I.amAble (2016-17) se hizo hincapié en el diseño y selección de fichas para hacer talleres (aunque también se realizaron talleres). En la segunda edición (2017-18) se puso un mayor énfasis en llevar los talleres a un mayor número de centros educativos. En la pasada edición (2018-19) se puso el acento en los procesos de evaluación. En esta cuarta edición (2019-20), se han seguido trabajando y puliendo todos esos aspectos, pero se ha priorizado la transformación de I.amAble en un proyecto de tipo aprendizaje-servicio, integrándolo en asignaturas formales, concretamente en Complementos de Física y Complementos de Química, del Máster en Formación de Profesorado, en la especialidad de Física y Química. Item The ribotoxin -sarcin can cleave the sarcin/ricin loop on late 60S pre-ribosomes(Nucleic Acids Research, 2020) Olombrada, Miriam; Peña, Cohue; Rodríguez Galán, Olga; Klingauf Nerurkar, Purnima; Portugal Calisto, Daniela; Oborská Oplová, Michaela; Altvater, Martin; Gavilanes, José G.; Martínez Del Pozo, Álvaro; Cruz, Jesús de la; García Ortega, Lucía; Govind Panse, VikramThe ribotoxin -sarcin belongs to a family of ribonucleases that cleave the sarcin/ricin loop (SRL), a critical functional rRNA element within the large ribosomal subunit (60S), thereby abolishing translation. Whether -sarcin targets the SRL only in mature 60S subunits remains unresolved. Here, we show that, in yeast, -sarcin can cleave SRLs within late 60S pre-ribosomes containing mature 25S rRNA but not nucleolar/nuclear 60S pre-ribosomes containing 27S pre-rRNA in vivo. Conditional expression of -sarcin is lethal, but does not impede early pre-rRNA processing, nuclear export and the cytoplasmic maturation of 60S pre-ribosomes. Thus, SRL-cleaved containing late 60S pre-ribosomes seem to escape cytoplasmic proofreading steps. Polysome analyses revealed that SRL-cleaved 60S ribosomal subunits form 80S initiation complexes, but fail to progress to the step of translation elongation. We suggest that the functional integrity of a -sarcin cleaved SRL might be assessed only during translation.Item Structural foundations of sticholysin functionality(BBA - Proteins and Proteomics, 2021) Palacios Ortega, Juan; García Linares, Sara; Rivera de Torre, Esperanza; Heras Márquez, Diego; Gavilanes, José G.; Peter, Slotte; Martínez Del Pozo, ÁlvaroActinoporins constitute a family of α pore-forming toxins produced by sea anemones. The soluble fold of these proteins consists of a β-sandwich flanked by two α-helices. Actinoporins exert their activity by specifically recognizing sphingomyelin at their target membranes. Once there, they penetrate the membrane with their N-terminal α-helices, a process that leads to the formation of cation-selective pores. These pores kill the target cells by provoking an osmotic shock on them. In this review, we examine the role and relevance of the structural features of actinoporins, down to the residue level. We look at the specific amino acids that play significant roles in the function of actinoporins and their fold. Particular emphasis is given to those residues that display a high degree of conservation across the actinoporin sequences known to date. In light of the latest findings in the field, the membrane requirements for pore formation, the effect of lipid composition, and the process of pore formation are also discussed.Item Evaluation of different approaches used to study membrane permeabilization by actinoporins on model lipid vesicles(BBA. Biomembranes, 2020) Palacios Ortega, Juan; Rivera de la Torre, Esperanza; Gavilanes, José G.; Slotte, J. Peter; Martínez Del Pozo, ÁlvaroRelease of aqueous contents from model lipid vesicles has been a standard procedure to evaluate pore formation efficiency by actinoporins, such as sticholysin II (StnII), for the last few decades. However, regardless of the probe of choice, the results reported that StnII action was never able to empty the vesicles completely. This was hard to explain if StnII pores were to be stable and always leaky for the probes used. To address this question, we have used a variety of probes, including rhodamine 6G or Tb3+, to test the permeability of StnII’s pores. Our results indicate that calcein was in fact too large to fit through StnII’s pores, and that the standard method in the field is actually reporting StnII-induced transient permeation of the membrane rather than the passage of solutes through the stable assembled pores. In order to evaluate the permeability of these structures, we used a dithionite-based assay, which showed that the final pores were in fact open. Thus, our results indicate that the stable actinoporins’ pores are open in spite of plateaued classic release curves. Besides the proper pore, the first stages of pore formation would inflict serious damage to living cells as well.Item Evaluation of different approaches used to study membrane permeabilization by actinoporins on model lipid vesicles(Biochimica et Biophysica Acta (BBA) - Biomembranes, 2020) Palacios Ortega, Juan; Rivera-de-Torre, Esperanza; Gavilanes, José G.; Slotte, J. Peter; Martínez Del Pozo, ÁlvaroRelease of aqueous contents from model lipid vesicles has been a standard procedure to evaluate pore formation efficiency by actinoporins, such as sticholysin II (StnII), for the last few decades. However, regardless of the probe of choice, the results reported that StnII action was never able to empty the vesicles completely. This was hard to explain if StnII pores were to be stable and always leaky for the probes used. To address this question, we have used a variety of probes, including rhodamine 6G or Tb3+, to test the permeability of StnII's pores. Our results indicate that calcein was in fact too large to fit through StnII's pores, and that the standard method in the field is actually reporting StnII-induced transient permeation of the membrane rather than the passage of solutes through the stable assembled pores. In order to evaluate the permeability of these structures, we used a dithionite-based assay, which showed that the final pores were in fact open. Thus, our results indicate that the stable actinoporins' pores are open in spite of plateaued classic release curves. Besides the proper pore, the first stages of pore formation would inflict serious damage to living cells as well.Item Sticholysin I–II oligomerization in the absence of membranes(FEBS Letters, 2022) García Linares, Sara; Amigot Sánchez, Rafael; García Montoya, Carmen; Alfonso, Carlos; Luque Ortega, Juan Román; Gavilanes, José G.; Martínez Del Pozo, Álvaro; Palacios Ortega, JuanSticholysins are pore-forming toxins produced by the sea anemone Stichodactyla helianthus. When they encounter a sphingomyelin-containing membrane, these proteins bind to it and oligomerize, a process that ends in pore formation. Mounting evidence indicates that StnII can favour the activity of StnI. Previous results have shown that these two isotoxins can oligomerize together. Furthermore, StnII appeared to potentiate the activity of StnI through the membrane-binding step of the process. Hence, isotoxin interaction should occur prior to membrane encounter. Here, we have used analytical ultracentrifugation to investigate the oligomerization of Stns in solution, both separately and together. Our results indicate that while StnI seems to be more prone to oligomerize in water solution than StnII, a small percentage of StnII in StnI–StnII mixtures promotes oligomerization.Item Structural and functional characterization of Sticholysin III: A newly discovered actinoporin within the venom of the sea anemone Stichodactyla helianthus.(Archives of Biochemistry and Biophysics, 2020) Rivera de Torre, Esperanza; Palacios Ortega, Juan; Garb, Jessica E.; Slotte, J. Peter; Gavilanes, José G.; Martínez Del Pozo, ÁlvaroActinoporins are a family of pore-forming toxins produced by sea anemones as part of their venomous cocktail. These proteins remain soluble and stably folded in aqueous solution, but when interacting with sphingomyelin-containing lipid membranes, they become integral oligomeric membrane structures that form a pore permeable to cations, which leads to cell death by osmotic shock. Actinoporins appear as multigenic families within the genome of sea anemones: several genes encoding very similar actinoporins are detected within the same species. The Caribbean Sea anemone Stichodactyla helianthus produces three actinoporins (sticholysins I, II and III; StnI, StnII and StnIII) that differ in their toxic potency. For example, StnII is about four-fold more effective than StnI against sheep erythrocytes in causing hemolysis, and both show synergy. However, StnIII, recently discovered in the S. helianthus transcriptome, has not been characterized so far. Here we describe StnIII’s spectroscopic and functional properties and show its potential to interact with the other Stns. StnIII seems to maintain the well-preserved fold of all actinoporins, characterized by a high content of β-sheet, but it is significantly less thermostable. Its functional characterization shows that the critical concentration needed to form active pores is higher than for either StnI or StnII, suggesting differences in behavior when oligomerizing on membrane surfaces. Our results show that StnIII is an interesting and unexpected piece in the puzzle of how this Caribbean Sea anemone species modulates its venomous activity.Item Functional and Structural Variation among Sticholysins, Pore-Forming Proteins from the Sea Anemone Stichodactyla helianthus(International journal of molecular sciences, 2020) Rivera de la Torre, Esperanza; Palacios Ortega, Juan; Slotte, J. Peter; Gavilanes, José G.; Martínez Del Pozo, Álvaro; García Linares, SaraVenoms constitute complex mixtures of many different molecules arising from evolution in processes driven by continuous prey–predator interactions. One of the most common compounds in these venomous cocktails are pore-forming proteins, a family of toxins whose activity relies on the disruption of the plasmatic membranes by forming pores. The venom of sea anemones, belonging to the oldest lineage of venomous animals, contains a large amount of a characteristic group of pore-forming proteins known as actinoporins. They bind specifically to sphingomyelin-containing membranes and suffer a conformational metamorphosis that drives them to make pores. This event usually leads cells to death by osmotic shock. Sticholysins are the actinoporins produced by Stichodactyla helianthus. Three different isotoxins are known: Sticholysins I, II, and III. They share very similar amino acid sequence and three-dimensional structure but display different behavior in terms of lytic activity and ability to interact with cholesterol, an important lipid component of vertebrate membranes. In addition, sticholysins can act in synergy when exerting their toxin action. The subtle, but important, molecular nuances that explain their different behavior are described and discussed throughout the text. Improving our knowledge about sticholysins behavior is important for eventually developing them into biotechnological tools.Item Oligomerization of Sticholysins from Förster Resonance Energy Transfer(Biochemistry, 2021) Palacios-Ortega, Juan; Rivera de la Torre, Esperanza; García-Linares, Sara; Gavilanes, José G.; Martínez Del Pozo, Álvaro; Slotte, J. PeterSticholysins are pore-forming toxins produced by sea anemones that are members of the actinoporin family. They exert their activity by forming pores on membranes, provided they have sphingomyelin. To assemble into pores, specific recognition, binding, and oligomerization are required. While recognition and binding have been extensively studied, delving into the oligomerization process and the stoichiometry of the pores has been more difficult. Here, we present evidence that these toxins are capable of oligomerizing in solution and suggesting that the interaction of sticholysin II (StnII) with its isoform sticholysin I (StnI) is stronger than that of StnI with itself. We also show that the stoichiometry of the final, thermodynamically stable StnI pores is, at least, heptameric. Furthermore, our results indicate that this association maintains its oligomerization number when StnII is included, indicating that the stoichiometry of StnII is also of that order, and not tetrameric, as previously thought. These results are compatible with the stoichiometry observed for the crystallized pore of FraC, another very similar actinoporin produced by a different sea anemone species. Our results also indicate that the stoichiometry of actinoporin pores in equilibrium is conserved regardless of the particular composition of a given pore ensemble, which we have shown for mixed sticholysin pores.