scholarly journals A novel yeast-based high-throughput method for the identification of protein palmitoylation inhibitors

Open Biology ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 200415
Author(s):  
Consuelo Coronel Arrechea ◽  
María Luz Giolito ◽  
Iris Alejandra García ◽  
Gastón Soria ◽  
Javier Valdez Taubas
Keyword(s):  
High Throughput ◽  
Protein S ◽  
Cell Physiology ◽  
Post Translational Modification ◽  
Lipid Molecule ◽  
Protein Palmitoylation ◽  
High Throughput Method ◽  
Thioester Bond ◽  
The Central Nervous System ◽  
Candidate Molecule

Protein S-acylation or palmitoylation is a widespread post-translational modification that consists of the addition of a lipid molecule to cysteine residues of proteins through a thioester bond. Palmitoylation and palmitoyltransferases (PATs) have been linked to several types of cancers, diseases of the central nervous system and many infectious diseases where pathogens use the host cell machinery to palmitoylate their effectors. Despite the central importance of palmitoylation in cell physiology and disease, progress in the field has been hampered by the lack of potent-specific inhibitors of palmitoylation in general, and of individual PATs in particular. Herein, we present a yeast-based method for the high-throughput identification of small molecules that inhibit protein palmitoylation. The system is based on a reporter gene that responds to the acylation status of a palmitoylation substrate fused to a transcription factor. The method can be applied to heterologous PATs such as human DHHC20, mouse DHHC21 and also a PAT from the parasite Giardia lamblia . As a proof-of-principle, we screened for molecules that inhibit the palmitoylation of Yck2, a substrate of the yeast PAT Akr1. We tested 3200 compounds and were able to identify a candidate molecule, supporting the validity of our method.

scholarly journals SwissPalm: Protein Palmitoylation database

F1000Research ◽  
2015 ◽  
Vol 4 ◽  
pp. 261 ◽  
Author(s):  
Mathieu Blanc ◽  
Fabrice David ◽  
Laurence Abrami ◽  
Daniel Migliozzi ◽  
Florence Armand ◽  
...  
Keyword(s):  
Protein S ◽  
Biological Processes ◽  
Post Translational Modification ◽  
Chemical Methods ◽  
Post Translational Modifications ◽  
P Protein ◽  
Multiple Alignments ◽  
Protein Palmitoylation ◽  
Recent Developments ◽  
Pathway Analyses

Protein S-palmitoylation is a reversible post-translational modification that regulates many key biological processes, although the full extent and functions of protein S-palmitoylation remain largely unexplored. Recent developments of new chemical methods have allowed the establishment of palmitoyl-proteomes of a variety of cell lines and tissues from different species.  As the amount of information generated by these high-throughput studies is increasing, the field requires centralization and comparison of this information. Here we present SwissPalm (http://swisspalm.epfl.ch), our open, comprehensive, manually curated resource to study protein S-palmitoylation. It currently encompasses more than 5000 S-palmitoylated protein hits from seven species, and contains more than 500 specific sites of S-palmitoylation. SwissPalm also provides curated information and filters that increase the confidence in true positive hits, and integrates predictions of S-palmitoylated cysteine scores, orthologs and isoform multiple alignments. Systems analysis of the palmitoyl-proteome screens indicate that 10% or more of the human proteome is susceptible to S-palmitoylation. Moreover, ontology and pathway analyses of the human palmitoyl-proteome reveal that key biological functions involve this reversible lipid modification. Comparative analysis finally shows a strong crosstalk between S-palmitoylation and other post-translational modifications. Through the compilation of data and continuous updates, SwissPalm will provide a powerful tool to unravel the global importance of protein S-palmitoylation.

A novel motif at the C-terminus of palmitoyltransferases is essential for Swf1 and Pfa3 function in vivo

2009 ◽  
Vol 419 (2) ◽  
pp. 301-308 ◽  
Author(s):  
Ayelén González Montoro ◽  
Rodrigo Quiroga ◽  
Hugo J. F. Maccioni ◽  
Javier Valdez Taubas
Keyword(s):  
Membrane Proteins ◽  
Post Translational Modification ◽  
Lipid Molecule ◽  
Additional Information ◽  
C Terminus ◽  
Protein Receptors ◽  
Thioester Bond ◽  
Function Relationship ◽  
Eukaryotic Organisms

S-acylation (commonly known as palmitoylation) is a widespread post-translational modification that consists of the addition of a lipid molecule to cysteine residues of a protein through a thioester bond. This modification is predominantly mediated by a family of proteins referred to as PATs (palmitoyltransferases). Most PATs are polytopic membrane proteins, with four to six transmembrane domains, a conserved DHHC motif and variable C-and N-terminal regions, that are probably responsible for conferring localization and substrate specificity. There is very little additional information on the structure–function relationship of PATs. Swf1 and Pfa3 are yeast members of the DHHC family of proteins. Swf1 is responsible for the S-acylation of several transmembrane SNAREs (soluble N-ethylmaleimide-sensitive fusion protein-attachment protein receptors) and other integral membrane proteins. Pfa3 is required for the palmitoylation of Vac8, a protein involved in vacuolar fusion. In the present study we describe a novel 16-amino-acid motif present at the cytosolic C-terminus of PATs, that is required for Swf1 and Pfa3 function in vivo. Within this motif, we have identified a single residue in Swf1, Tyr323, as essential for function, and this is correlated with lack of palmitoylation of Tlg1, a SNARE that is a substrate of Swf1. The equivalent mutation in Pfa3 also affects its function. These mutations are the first phenotype-affecting mutations uncovered that do not lie within the DHHC domain, for these or any other PATs. The motif is conserved in 70% of PATs from all eukaryotic organisms analysed, and may have once been present in all PATs. We have named this motif PaCCT (‘Palmitoyltransferase Conserved C-Terminus’).

A Fluorescent High Throughput Method To Identify Potential Skin Sensitizers

Planta Medica ◽  
2016 ◽  
Vol 82 (05) ◽  
Author(s):  
C Avonto ◽  
AG Chittiboyina ◽  
D Rua ◽  
IA Khan
Keyword(s):  
High Throughput ◽  
High Throughput Method

scholarly journals High Throughput Centrifugal Electrospinning of Polyacrylonitrile Nanofibers for Carbon Fiber Nonwovens

Polymers ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1313
Author(s):  
Andreas Hoffmann ◽  
Alexander J. C. Kuehne
Keyword(s):  
High Throughput ◽  
Carbon Nanofiber ◽  
Fiber Diameter ◽  
Carbon Precursor ◽  
Production Methods ◽  
High Throughput Method ◽  
Radial Forces ◽  
Fiber Deposition ◽  
The Impact ◽  
Pan Fibers

Carbon nanofiber nonwovens are promising materials for electrode or filtration applications; however, their utilization is obviated by a lack of high throughput production methods. In this study, we utilize a highly effective high-throughput method for the fabrication of polyacrylonitrile (PAN) nanofibers as a nonwoven on a dedicated substrate. The method employs rotational-, air pressure- and electrostatic forces to produce fibers from the inner edge of a rotating bell towards a flat collector. We investigate the impact of all above-mentioned forces on the fiber diameter, morphology, and bundling of the carbon-precursor PAN fibers. The interplay of radial forces with collector-facing forces has an influence on the uniformity of fiber deposition. Finally, the obtained PAN nanofibers are converted to carbon nonwovens by thermal treatment.

scholarly journals A neural network-based algorithm for high-throughput characterisation of viscoelastic properties of flowing microcapsules

Soft Matter ◽  
2021 ◽  
Author(s):  
Tao Lin ◽  
Zhen Wang ◽  
Wen Wang ◽  
Yi Sui
Keyword(s):  
Neural Network ◽  
High Throughput ◽  
Viscoelastic Properties ◽  
Dynamic Deformation ◽  
Membrane Elasticity ◽  
Hybrid Neural Network ◽  
High Throughput Method

We have developed a high-throughput method, by combining a hybrid neural network with a mechanistic capsule model, to predict membrane elasticity and viscosity of microcapsules from their dynamic deformation in a branched microchannel.

scholarly journals Development of a High-Throughput Method to Study the Inhibitory Effect of Phytochemicals on Trimethylamine Formation

Nutrients ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 1466
Author(s):  
Lisard Iglesias-Carres ◽  
Lauren A. Essenmacher ◽  
Kathryn C. Racine ◽  
Andrew P. Neilson
Keyword(s):  
Chlorogenic Acid ◽  
Gallic Acid ◽  
High Throughput ◽  
Anaerobic Fermentation ◽  
Inhibitory Effect ◽  
Healthy Human ◽  
High Throughput Method ◽  
Lyase Activity ◽  
Inhibitory Potential ◽  
Potential Maximum

Choline is metabolized by the gut microbiota into trimethylamine (TMA), the precursor of pro-atherosclerotic molecule trimethylamine N-oxide (TMAO). A reduction in TMA formation has shown cardioprotective effects, and some phytochemicals may reduce TMA formation. This study aimed to develop an optimized, high-throughput anaerobic fermentation methodology to study the inhibition of choline microbial metabolism into TMA by phenolic compounds with healthy human fecal starter. Optimal fermentation conditions were: 20% fecal slurry (1:10 in PBS), 100 µM choline, and 12 h fermentation. Additionally, 10 mM of 3,3-dimethyl-1-butanol (DMB) was defined as a positive TMA production inhibitor, achieving a ~50% reduction in TMA production. Gallic acid and chlorogenic acid reported higher TMA inhibitory potential (maximum of 80–90% TMA production inhibition), with IC50 around 5 mM. Neither DMB nor gallic acid or chlorogenic acid reduced TMA production through cytotoxic effects, indicating mechanisms such as altered TMA-lyase activity or expression.

scholarly journals High-Throughput Chlorophyll and Carotenoid Profiling Reveals Positive Associations with Sugar and Apocarotenoid Volatile Content in Fruits of Tomato Varieties in Modern and Wild Accessions

Metabolites ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 398
Author(s):  
Yusuke Aono ◽  
Yonathan Asikin ◽  
Ning Wang ◽  
Denise Tieman ◽  
Harry Klee ◽  
...  
Keyword(s):  
High Throughput ◽  
Fruit Color ◽  
Wild Relatives ◽  
Aroma Compounds ◽  
Soluble Solids ◽  
Sugar Production ◽  
Volatile Content ◽  
Factors Influencing ◽  
High Throughput Method ◽  
Wild Accessions

Flavor and nutritional quality has been negatively impacted during the course of domestication and improvement of the cultivated tomato (Solanum lycopersicum). Recent emphasis on consumers has emphasized breeding strategies that focus on flavor-associated chemicals, including sugars, acids, and aroma compounds. Carotenoids indirectly affect flavor as precursors of aroma compounds, while chlorophylls contribute to sugar production through photosynthesis. However, the relationships between these pigments and flavor content are still unclear. In this study, we developed a simple and high-throughput method to quantify chlorophylls and carotenoids. This method was applied to over one hundred tomato varieties, including S. lycopersicum and its wild relatives (S. l. var. cerasiforme and S. pimpinellifolium), for quantification of these pigments in fruits. The results obtained by integrating data of the pigments, soluble solids, sugars, and aroma compounds indicate that (i) chlorophyll-abundant varieties have relatively higher sugar accumulations and (ii) prolycopene is associated with an abundance of linear carotenoid-derived aroma compounds in one of the orange-fruited varieties, “Dixie Golden Giant”. Our results suggest the importance of these pigments not only as components of fruit color but also as factors influencing flavor traits, such as sugars and aroma.

scholarly journals Feedback Regulation of O-GlcNAc Transferase through Translation Control to Maintain Intracellular O-GlcNAc Homeostasis

2021 ◽  
Vol 22 (7) ◽  
pp. 3463
Author(s):  
Chia-Hung Lin ◽  
Chen-Chung Liao ◽  
Mei-Yu Chen ◽  
Teh-Ying Chou
Keyword(s):  
Molecular Mechanisms ◽  
Mrna Level ◽  
Feedback Regulation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Cell Physiology ◽  
Hydroxyl Groups ◽  
Post Translational Modification ◽  
Translation Control ◽  
Glcnac Transferase

Protein O-GlcNAcylation is a dynamic post-translational modification involving the attachment of N-acetylglucosamine (GlcNAc) to the hydroxyl groups of Ser/Thr residues on numerous nucleocytoplasmic proteins. Two enzymes are responsible for O-GlcNAc cycling on substrate proteins: O-GlcNAc transferase (OGT) catalyzes the addition while O-GlcNAcase (OGA) helps the removal of GlcNAc. O-GlcNAcylation modifies protein functions; therefore, dysregulation of O-GlcNAcylation affects cell physiology and contributes to pathogenesis. To maintain homeostasis of cellular O-GlcNAcylation, there exists feedback regulation of OGT and OGA expression responding to fluctuations of O-GlcNAc levels; yet, little is known about the molecular mechanisms involved. In this study, we investigated the O-GlcNAc-feedback regulation of OGT and OGA expression in lung cancer cells. Results suggest that, upon alterations in O-GlcNAcylation, the regulation of OGA expression occurs at the mRNA level and likely involves epigenetic mechanisms, while modulation of OGT expression is through translation control. Further analyses revealed that the eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) contributes to the downregulation of OGT induced by hyper-O-GlcNAcylation; the S5A/S6A O-GlcNAcylation-site mutant of 4E-BP1 cannot support this regulation, suggesting an important role of O-GlcNAcylation. The results provide additional insight into the molecular mechanisms through which cells may fine-tune intracellular O-GlcNAc levels to maintain homeostasis.

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