Examinando por Autor "Rojas, Maximiliano"
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Ítem Different classes of antidepressants inhibit the rat α7 nicotinic acetylcholine receptor by interacting within the ion channel: A functional and structural study(MDPI, 2021-02) Duarte, Yorley; Rojas, Maximiliano; Canan, Jonathan; Pérez, Edwin G.; González-Nilo, Fernando; García-Colunga, JesúsSeveral antidepressants inhibit nicotinic acetylcholine receptors (nAChRs) in a noncompetitive and voltage-dependent fashion. Here, we asked whether antidepressants with a different structure and pharmacological pro le modulate the rat α7 nAChR through a similar mechanism by interacting within the ion-channel. We applied electrophysiological (recording of the ion current elicited by choline, ICh, which activates α7 nAChRs from rat CA1 hippocampal interneurons) and in silico approaches (homology modeling of the rat α7 nAChR, molecular docking, molecular dynamics simulations, and binding free energy calculations). The antidepressants inhibited IChwith the order: Norfluoxetine ~ mirtazapine ~ imipramine < bupropion ~ fluoxetine ~ venlafaxine ~ escitalopram. The constructed homology model of the rat α7 nAChR resulted in the extracellular vestibule and the channel pore is highly negatively charged, which facilitates the permeation of cations and the entrance of the protonated form of antidepressants. Molecular docking and molecular dynamics simulations were carried out within the ion-channel of the α7 nAChR, revealing that the antidepressants adopt poses along the receptor channel, with slightly different binding-free energy values. Furthermore, the inhibition of ICh and free energy values for each antidepressant-receptor complex were highly correlated. Thus, the α7 nAChR is negatively modulated by a variety of antidepressants interacting in the ion-channel. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.Ítem Endogenous pannexin1 channels form functional intercellular cell-cell channels with characteristic voltage-dependent properties(National Academy of Sciences, 2022-05-03) Palacios-Prado, Nicolas; Soto, Paola A.; Lopez, Ximena; Choi, Eun Ju; Marquez-Miranda, Valeria; Rojas, Maximiliano; Duarte, Yorley; Lee, Jinu; Gonzalez-Nilo, Fernando D.; Saez, Juan C.The occurrence of intercellular channels formed by pannexin1 has been challenged for more than a decade. Here, we provide an electrophysiological characterization of exogenous human pannexin1 (hPanx1) cell-cell channels expressed in HeLa cells knocked out for connexin45. The observed hPanx1 cell-cell channels show two phenotypes: O-state and S-state. The former displayed low transjunctional voltage (Vj) sensitivity and singlechannel conductance of ∼175 pS, with a substate of ∼35 pS; the latter showed a peculiar dynamic asymmetry in Vj dependence and single-channel conductance identical to the substate conductance of the O-state. S-state hPanx1 cell-cell channels were also identified between TC620 cells, a human oligodendroglioma cell line that endogenously expresses hPanx1. In these cells, dye and electrical coupling increased with temperature and were strongly reduced after hPanx1 expression was knocked down by small interfering RNA or inhibited with Panx1 mimetic inhibitory peptide. Moreover, cell-cell coupling was augmented when hPanx1 levels were increased with a doxycycline-inducible expression system. Application of octanol, a connexin gap junction (GJ) channel inhibitor, was not sufficient to block electrical coupling between HeLa KO Cx45-hPanx1 or TC620 cell pairs. In silico studies suggest that several arginine residues inside the channel pore may be neutralized by hydrophobic interactions, allowing the passage of DAPI, consistent with dye coupling observed between TC620 cells. These findings demonstrate that endogenously expressed hPanx1 forms intercellular cell-cell channels and their unique properties resemble those described in innexin-based GJ channels. Since Panx1 is ubiquitously expressed, finding conditions to recognize Panx1 cell-cell channels in different cell types might require special attention. © 2022 National Academy of Sciences. All rights reserved.Ítem Increased Absorption of Thyroxine in a Murine Model of Hypothyroidism Using Water/CO2 Nanobubbles(Multidisciplinary Digital Publishing Institute (MDPI), 2024-06) Opazo, Maria Cecilia; Yañez, Osvaldo; Márquez-Miranda, Valeria; Santos, Johana; Rojas, Maximiliano; Araya-Durán, Ingrid; Aguayo, Daniel; Leal, Matías; Duarte, Yorley; Kohanoff, Jorge; González-Nilo, Fernando D.Thyroxine (T4) is a drug extensively utilized for the treatment of hypothyroidism. However, the oral absorption of T4 presents certain limitations. This research investigates the efficacy of CO2 nanobubbles in water as a potential oral carrier for T4 administration to C57BL/6 hypothyroid mice. Following 18 h of fasting, the formulation was administered to the mice, demonstrating that the combination of CO2 nanobubbles and T4 enhanced the drug’s absorption in blood serum by approximately 40%. To comprehend this observation at a molecular level, we explored the interaction mechanism through which T4 engages with the CO2 nanobubbles, employing molecular simulations, semi-empirical quantum mechanics, and PMF calculations. Our simulations revealed a high affinity of T4 for the water–gas interface, driven by additive interactions between the hydrophobic region of T4 and the gas phase and electrostatic interactions of the polar groups of T4 with water at the water–gas interface. Concurrently, we observed that at the water–gas interface, the cluster of T4 formed in the water region disassembles, contributing to the drug’s bioavailability. Furthermore, we examined how the gas within the nanobubbles aids in facilitating the drug’s translocation through cell membranes. This research contributes to a deeper understanding of the role of CO2 nanobubbles in drug absorption and subsequent release into the bloodstream. The findings suggest that utilizing CO2 nanobubbles could enhance T4 bioavailability and cell permeability, leading to more efficient transport into cells. Additional research opens the possibility of employing lower concentrations of this class of drugs, thereby potentially reducing the associated side effects due to poor absorption.Ítem The Emergence of New Catalytic Abilities in an Endoxylanase from Family GH10 by Removing an Intrinsically Disordered Region(MDPI, 2022-02-02) Gil-Durán, Carlos; Sepúlveda, Romina V.; Rojas, Maximiliano; Castro-Fernández, Víctor; Guixé, Victoria; Vaca, Inmaculada; Levicán, Gloria; González-Nilo, Fernando D.; Ravanal, María-Cristina; Chávez, RenatoEndoxylanases belonging to family 10 of the glycoside hydrolases (GH10) are versatile in the use of different substrates. Thus, an understanding of the molecular mechanisms underlying substrate specificities could be very useful in the engineering of GH10 endoxylanases for biotechno-logical purposes. Herein, we analyzed XynA, an endoxylanase that contains a (β/α)8-barrel domain and an intrinsically disordered region (IDR) of 29 amino acids at its amino end. Enzyme activity assays revealed that the elimination of the IDR resulted in a mutant enzyme (XynA∆29) in which two new activities emerged: the ability to release xylose from xylan, and the ability to hydrolyze p-nitrophenyl-β-D-xylopyranoside (pNPXyl), a substrate that wild-type enzyme cannot hydrolyze. Circular dichroism and tryptophan fluorescence quenching by acrylamide showed changes in secondary structure and increased flexibility of XynA∆29. Molecular dynamics simulations revealed that the emergence of the pNPXyl-hydrolyzing activity correlated with a dynamic behavior not previously observed in GH10 endoxylanases: a hinge-bending motion of two symmetric regions within the (β/α)8-barrel domain, whose hinge point is the active cleft. The hinge-bending motion is more intense in XynA∆29 than in XynA and promotes the formation of a wider active site that allows the accommodation and hydrolysis of pNPXyl. Our results open new avenues for the study of the relationship between IDRs, dynamics and activity of endoxylanases, and other enzymes containing (β/α)8-barrel domain.Ítem Virtual Screening of Plant Volatile Compounds Reveals a High Affinity of Hylamorpha elegans (Coleoptera: Scarabaeidae) Odorant-Binding Proteins for Sesquiterpenes From Its Native Host(Oxford Univ Press, 2016) González-González, Angélica; Palma-Millanao, Rubén; Yáñez, Osvaldo; Rojas, Maximiliano; Mutis, Ana; Venthur, Herbert; Quiroz, Andrés; Ramírez, Claudio C.Hylamorpha elegans (Burmeister) is a native Chilean scarab beetle considered to be a relevant agricultural pest to pasture and cereal and small fruit crops. Because of their cryptic habits, control with conventional methods is difficult; therefore, alternative and environmentally friendly control strategies are highly desirable. The study of proteins that participate in the recognition of odorants, such as odorant-binding proteins (OBPs), offers interesting opportunities to identify new compounds with the potential to modify pest behavior and computational screening of compounds, which is commonly used in drug discovery, may help to accelerate the discovery of new semiochemicals. Here, we report the discovery of four OBPs in H. elegans as well as six new volatiles released by its native host Nothofagus obliqua (Mirbel). Molecular docking performed between OBPs and new and previously reported volatiles from N. obliqua revealed the best binding energy values for sesquiterpenic compounds. Despite remarkable divergence at the amino acid level, three of the four OBPs evaluated exhibited the best interaction energy for the same ligands. Molecular dynamics investigation reinforced the importance of sesquiterpenes, showing that hydrophobic residues of the OBPs interacted most frequently with the tested ligands, and binding free energy calculations demonstrated van der Waals and hydrophobic interactions to be the most important. Altogether, the results suggest that sesquiterpenes are interesting candidates for in vitro and in vivo assays to assess their potential application in pest management strategies.