Frontispiece: The Impact of Heteromultivalency in Lectin Recognition and Glycosidase Inhibition: An Integrated Mechanistic Study

Background: Potentially novel regulators of early human germline development have been identified recently, including SOX15 and SOX17, both of which show specific expression in human primordial germ cells. SOX17 is now known to be a critical specifier of human germ cell identity. There have been suggestions, as yet without evidence, that SOX15 might also play a prominent role. The early human germline is inaccessible for direct study, but an in vitro model of human primordial germ cell-like cell (hPGCLC) specification from human embryonic stem cells (hESCs) has been developed. This enables mechanistic study of human germ cell specification using genetic tools to manipulate the levels of SOX15 and SOX17 proteins to explore their roles in hPGCLC specification. Methods: SOX15 and SOX17 proteins were depleted during hPGCLC specification from hESCs using the auxin-inducible degron system, combined with a fluorescent reporter for tracking protein levels. Additionally, SOX15 protein was overexpressed using the ProteoTuner system. Protein-level expression changes were confirmed by immunofluorescence. The impact on hPGCLC specification efficiency was determined by flow cytometry at various time points. qPCR experiments were performed to determine some transcriptional effects of SOX15 perturbations. Results: We observed specific SOX15 expression in hPGCLCs by using immunofluorescence and flow cytometry analysis. Depletion of SOX15 had no significant effect on hPGCLC specification efficiency on day 4 after induction, but there was a significant and progressive decrease in hPGCLCs on days 6 and 8. By contrast, depletion of SOX17 completely abrogated hPGCLC specification. Furthermore, SOX15 overexpression resulted in a significant increase in hPGCLC fraction on day 8. qPCR analysis revealed a possible role for the germ cell and pluripotency regulator PRDM14 in compensating for changes to SOX15 protein levels. Conclusions: SOX17 is essential for hPGCLC specification, yet SOX15 is dispensable. However, SOX15 may have a role in maintaining germ cell identity.

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Studies on the Kinetics of Doxazosin Degradation in Simulated Environmental Conditions and Selected Advanced Oxidation Processes

Water ◽

10.3390/w11051001 ◽

2019 ◽

Vol 11 (5) ◽

pp. 1001 ◽

Cited By ~ 3

Author(s):

Joanna Karpinska ◽

Aneta Sokol ◽

Jolanta Koldys ◽

Artur Ratkiewicz

Keyword(s):

Environmental Conditions ◽

Density Functional ◽

Natural Waters ◽

Basic Medium ◽

Mechanistic Study ◽

Aqueous Environment ◽

Functional Theory ◽

Short Time ◽

The Impact ◽

Kinetics Of

The photochemical behavior of doxazosin (DOX) in simulated environmental conditions using natural waters taken from local rivers as a solvent was studied. The chemical characteristics of applied waters was done and a correlation analysis was used to explain the impact of individual parameters of matrix on the rate of the DOX degradation. It was stated that DOX is a photoliable compound in an aqueous environment. Its degradation is promoted by basic medium, presence of environmentally important ions such as Cl−, NO3−, SO42− and organic matter. The kinetics of DOX reactions with OH− and SO4− radicals were examined individually. The UV/H2O2, classical Fenton and photo-Fenton processes, were applied for the generation of hydroxyl radicals while the UV/VIS:Fe2(SO4)3:Na2SO2 system was employed for production of SO4− radicals. The obtained results pointed that photo-Fenton, as well as UV/VIS:Fe2(SO4)3:Na2SO2, are very reactive in ratio to DOX, leading to its complete degradation in a short time. A quantitative density functional theory (DFT) mechanistic study was carried out in order to explain the molecular mechanism of DOX degradation using the GAUSSIAN 09 program.

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A Mechanistic Study on Gold(I)-Catalyzed Cyclization of Propargylic Amide: Revealing the Impact of Expanded-Ring N-Heterocyclic Carbenes

Catalysis Science & Technology ◽

10.1039/d1cy01617b ◽

2021 ◽

Author(s):

Yumiao Ma ◽

Hafiz Saqib Ali ◽

Aqeel A. Hussein

Keyword(s):

Density Functional Theory ◽

Density Functional ◽

Heterocyclic Carbenes ◽

Mechanistic Study ◽

Functional Theory ◽

The Impact

The interest in expanded-ring N-heterocyclic carbenes (ER-NHCs) has been recently given a noticeable attention, especially with the Au(I)-catalyzed activation of alkynes. Herein, we report density functional theory (DFT) investigations on...

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Modeling the Alkaline Hydrolysis of Diaryl Sulfate Diesters: A Mechanistic Study

10.26434/chemrxiv.11874072.v1 ◽

2020 ◽

Author(s):

Klaudia Szeler ◽

Nicholas Williams ◽

Alvan C. Hengge ◽

Shina Caroline Lynn Kamerlin

Keyword(s):

Alkaline Hydrolysis ◽

Computational Study ◽

Transition States ◽

Building Blocks ◽

Ester Hydrolysis ◽

Mechanistic Study ◽

Phosphate Ester ◽

Sulfate Ester ◽

The Impact ◽

Hydrolysis Of

<div> <div> <div> <p>Phosphate and sulfate esters have important roles as biological building blocks and in regulating cellular processes. However, while there has been substantial experimental and computational investigation of the mechanisms and the transition states involved in phosphate ester hydrolysis, there is far less (in particular computational) work on sulfate ester hydrolysis. Here, we report a detailed computational study of the alkaline hydrolysis of diaryl sulfate diesters, using different DFT functionals and both pure implicit solvation as well as mixed implicit/explicit solvation with varying numbers of explicit water molecules. We consider both the impact of how the system is modeled on computed linear free energy relationships (LFER) and the nature of the transition states. Although our calculations consistently underestimate the absolute activation free energies, we obtain good agreement with experimental LFER data when using pure implicit solvent, and excellent agreement with experimental kinetic isotope effects for all models used. Our calculations suggest that the hydrolysis of sulfate diesters proceeds through loose transition states, with minimal bond formation to the nucleophile and with bond cleavage to the leaving group already initiated. Comparison to prior work indicates that these transition states are similar in nature to those of analogous reactions such as the alkaline hydrolysis of neutral arylsulfonate monoesters or charged phosphate diesters and fluorophosphates. Obtaining more detailed insight into the transition states involved assists in understanding the selectivity of enzymes that hydrolyze these reactions; however, this work also highlights the methodological challenges involved in reliably modeling sulfate ester hydrolysis. </p> </div> </div> </div>

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Design–functionality relationships for adhesion/growth-regulatory galectins

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1813515116 ◽

2019 ◽

Vol 116 (8) ◽

pp. 2837-2842 ◽

Cited By ~ 24

Author(s):

Anna-Kristin Ludwig ◽

Malwina Michalak ◽

Qi Xiao ◽

Ulrich Gilles ◽

Francisco J. Medrano ◽

...

Keyword(s):

Therapeutic Potential ◽

Growth Inhibitor ◽

Modular Architecture ◽

The Family ◽

Specific Contact ◽

Galectin 3 ◽

Marked Selectivity ◽

Functional Antagonism ◽

The Impact ◽

Lectin Recognition

Glycan-lectin recognition is assumed to elicit its broad range of (patho)physiological functions via a combination of specific contact formation with generation of complexes of distinct signal-triggering topology on biomembranes. Faced with the challenge to understand why evolution has led to three particular modes of modular architecture for adhesion/growth-regulatory galectins in vertebrates, here we introduce protein engineering to enable design switches. The impact of changes is measured in assays on cell growth and on bridging fully synthetic nanovesicles (glycodendrimersomes) with a chemically programmable surface. Using the example of homodimeric galectin-1 and monomeric galectin-3, the mutual design conversion caused qualitative differences, i.e., from bridging effector to antagonist/from antagonist to growth inhibitor and vice versa. In addition to attaining proof-of-principle evidence for the hypothesis that chimera-type galectin-3 design makes functional antagonism possible, we underscore the value of versatile surface programming with a derivative of the pan-galectin ligand lactose. Aggregation assays with N,N′-diacetyllactosamine establishing a parasite-like surface signature revealed marked selectivity among the family of galectins and bridging potency of homodimers. These findings provide fundamental insights into design-functionality relationships of galectins. Moreover, our strategy generates the tools to identify biofunctional lattice formation on biomembranes and galectin-reagents with therapeutic potential.

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Bacteria Lectin Recognition Towards Fucose Binding Motifs Highlights the Impact of Presenting Mucin Core Glycopeptides

10.33774/chemrxiv-2021-79qhk ◽

2021 ◽

Author(s):

Sandra Behren ◽

Jin Yu ◽

Christian Pett ◽

Manuel Shorlemer ◽

Viktoria Heine ◽

...

Keyword(s):

Binding Motifs ◽

The Impact ◽

Lectin Recognition

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Modeling the Alkaline Hydrolysis of Diaryl Sulfate Diesters: A Mechanistic Study

10.26434/chemrxiv.11874072 ◽

2020 ◽

Cited By ~ 1

Author(s):

Klaudia Szeler ◽

Nicholas Williams ◽

Alvan C. Hengge ◽

Shina Caroline Lynn Kamerlin

Keyword(s):

Alkaline Hydrolysis ◽

Computational Study ◽

Transition States ◽

Building Blocks ◽

Ester Hydrolysis ◽

Mechanistic Study ◽

Phosphate Ester ◽

Sulfate Ester ◽

The Impact ◽

Hydrolysis Of

<div> <div> <div> <p>Phosphate and sulfate esters have important roles as biological building blocks and in regulating cellular processes. However, while there has been substantial experimental and computational investigation of the mechanisms and the transition states involved in phosphate ester hydrolysis, there is far less (in particular computational) work on sulfate ester hydrolysis. Here, we report a detailed computational study of the alkaline hydrolysis of diaryl sulfate diesters, using different DFT functionals and both pure implicit solvation as well as mixed implicit/explicit solvation with varying numbers of explicit water molecules. We consider both the impact of how the system is modeled on computed linear free energy relationships (LFER) and the nature of the transition states. Although our calculations consistently underestimate the absolute activation free energies, we obtain good agreement with experimental LFER data when using pure implicit solvent, and excellent agreement with experimental kinetic isotope effects for all models used. Our calculations suggest that the hydrolysis of sulfate diesters proceeds through loose transition states, with minimal bond formation to the nucleophile and with bond cleavage to the leaving group already initiated. Comparison to prior work indicates that these transition states are similar in nature to those of analogous reactions such as the alkaline hydrolysis of neutral arylsulfonate monoesters or charged phosphate diesters and fluorophosphates. Obtaining more detailed insight into the transition states involved assists in understanding the selectivity of enzymes that hydrolyze these reactions; however, this work also highlights the methodological challenges involved in reliably modeling sulfate ester hydrolysis. </p> </div> </div> </div>

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Nanoemulsion adjuvantation strategy of tumor-associated antigen therapy rephrases mucosal and immunotherapeutic signatures following intranasal vaccination

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-001022 ◽

2020 ◽

Vol 8 (2) ◽

pp. e001022

Author(s):

Chung-Hsiung Huang ◽

Chiung-Yi Huang ◽

Hui-Min Ho ◽

Ching-Hung Lee ◽

Pang-Ti Lai ◽

...

Keyword(s):

Cellular Immunity ◽

Broad Spectrum ◽

Mechanistic Study ◽

Adjuvant Activity ◽

Mucosal Membrane ◽

Mucosal Tissues ◽

Cell Immunity ◽

Tumor Associated Antigen ◽

The Impact

BackgroundEmulsion adjuvants are a potent tool for effective vaccination; however, the size matters on mucosal signatures and the mechanism of action following intranasal vaccination remains unclear. Here, we launch a mechanistic study to address how mucosal membrane interacts with nanoemulsion of a well-defined size at cellular level and to elucidate the impact of size on tumor-associated antigen therapy.MethodsThe squalene-based emulsified particles at the submicron/nanoscale could be elaborated by homogenization/extrusion. The mucosal signatures following intranasal delivery in mice were evaluated by combining whole-mouse genome microarray and immunohistochemical analysis. The immunological signatures were tested by assessing their ability to influence the transportation of a model antigen ovalbumin (OVA) across nasal mucosal membranes and drive cellular immunity in vivo. Finally, the cancer immunotherapeutic efficacy is monitored by assessing tumor-associated antigen models consisting of OVA protein and tumor cells expressing OVA epitope.ResultsUniform structures with ~200 nm in size induce the emergence of membranous epithelial cells and natural killer cells in nasal mucosal tissues, facilitate the delivery of protein antigen across the nasal mucosal membrane and drive broad-spectrum antigen-specific T-cell immunity in nasal mucosal tissues as well as in the spleen. Further, intranasal vaccination of the nanoemulsion could assist the antigen to generate potent antigen-specific CD8+ cytotoxic T-lymphocyte response. When combined with immunotherapeutic models, such an effective antigen-specific cytotoxic activity allowed the tumor-bearing mice to reach up to 50% survival 40 days after tumor inoculation; moreover, the optimal formulation significantly attenuated lung metastasis.ConclusionsIn the absence of any immunostimulator, only 0.1% content of squalene-based nanoemulsion could rephrase the mucosal signatures following intranasal vaccination and induce broad-spectrum antigen-specific cellular immunity, thereby improving the efficacy of tumor-associated antigen therapy against in situ and metastatic tumors. These results provide critical mechanistic insights into the adjuvant activity of nanoemulsion and give directions for the design and optimization of mucosal delivery for vaccine and immunotherapy.

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