The mammalian two-hybrid system as a powerful tool for high-throughput drug screening

Patrício, Daniela; Fardilha, Margarida

doi:10.1016/j.drudis.2020.01.022

Cited by 4 publications

(5 citation statements)

References 57 publications

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“…Membrane two-hybrid assays developed in yeast and mammals are a less popular but also used technique to analyze the interaction of proteins with cell membranes. Their advantage is the ease of use in high-throughput screening tests [ 76 , 77 ]. Photoactivatable or DNA-based lipid probes are also of great importance in screening.…”

Section: Research Methods Used To Analyze Interactions With Cell Membranementioning

confidence: 99%

Interactions of Amyloid-β with Membrane Proteins

Wiatrak

Piasny

Kuźniarski

et al. 2021

IJMS

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In developing and developed countries, an increasing elderly population is observed. This affects the growing percentage of people struggling with neurodegenerative diseases, including Alzheimer’s disease. Nevertheless, the pathomechanism of this disease is still unknown. This contributes to problems with early diagnosis of the disease as well as with treatment. One of the most popular hypotheses of Alzheimer’s disease is related to the pathological deposition of amyloid-β (Aβ) in the brain of ill people. In this paper, we discuss issues related to Aβ and its relationship in the development of Alzheimer’s disease. The structure of Aβ and its interaction with the cell membrane are discussed. Not only do the extracellular plaques affect nerve cells, but other forms of this peptide as well.

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Section: Research Methods Used To Analyze Interactions With Cell Membranementioning

confidence: 99%

Interactions of Amyloid-β with Membrane Proteins

Wiatrak

Piasny

Kuźniarski

et al. 2021

IJMS

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“…Principles of the Y2H technology have also been adapted to mammalian systems, which circumvent some of the aforementioned drawbacks of Y2H. 881 Perhaps the most commonly applied method of detecting and validating PPIs in vitro is affinity purification (AP, also known as affinity chromatography) of co-immunoprecipitation (Co-IP) either coupled with SDS-PAGE/immunoblotting or mass spectrometry to determine the identity of interacting proteins. AP typically relies on the isolation of a transgenic POI by an epitope tag (using epitope-specific matrixconjugated proteins (antibodies or epitope-binding proteins)), while Co-IP harnesses specific antibodies directly targeting the POI.…”

Section: Validation Of Protein−protein Interactionsmentioning

confidence: 99%

“…when probing PPI of mammalian proteins in Y2H). Principles of the Y2H technology have also been adapted to mammalian systems, which circumvent some of the aforementioned drawbacks of Y2H …”

Section: Orthogonal Validationmentioning

confidence: 99%

Comprehensive Overview of Bottom-Up Proteomics Using Mass Spectrometry

Jiang,

Rex,

Schuster

et al. 2024

ACS Meas. Sci. Au

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Proteomics is the large scale study of protein structure and function from biological systems through protein identification and quantification. “Shotgun proteomics” or “bottom-up proteomics” is the prevailing strategy, in which proteins are hydrolyzed into peptides that are analyzed by mass spectrometry. Proteomics studies can be applied to diverse studies ranging from simple protein identification to studies of proteoforms, protein-protein interactions, protein structural alterations, absolute and relative protein quantification, post-translational modifications, and protein stability. To enable this range of different experiments, there are diverse strategies for proteome analysis. The nuances of how proteomic workflows differ may be challenging to understand for new practitioners. Here, we provide a comprehensive overview of different proteomics methods. We cover from biochemistry basics and protein extraction to biological interpretation and orthogonal validation. We expect this Review will serve as a handbook for researchers who are new to the field of bottom-up proteomics.

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“…Similarly, interactions of amyloid proteins can easily mislead the study, where interactions can result from the physical properties of the protein and not a specific interaction [19]. Various modifications of the classical Y2H system have been studied to elucidate the underlying interactions of the proteins (reviewed in [19,25,26]). Meanwhile, outcomes of the two hybrid analyses should be further evaluated by independent methods such as cross-linking experiments using materials from the original host.…”

Section: Yeast Two-hybrid (Y2h) Systemmentioning

confidence: 99%

The Use of Yeast in Biosensing

Dhakal

Macreadie

2022

Microorganisms

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Yeast has been used as a model for several diseases as it is the simplest unicellular eukaryote, safe and easy to culture and harbors most of the fundamental processes that are present in almost all higher eukaryotes, including humans. From understanding the pathogenesis of disease to drug discovery studies, yeast has served as an important biosensor. It is not only due to the conservation of genetics, amenable modification of its genome and easily accessible analytical methods, but also some characteristic features such as its ability to survive with defective mitochondria, making it a highly flexible microbe for designing whole-cell biosensing systems. The aim of this review is to report on how yeasts have been utilized as biosensors, reporting on responses to various stimuli.

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The mammalian two-hybrid system as a powerful tool for high-throughput drug screening

Cited by 4 publications

References 57 publications

Interactions of Amyloid-β with Membrane Proteins

Interactions of Amyloid-β with Membrane Proteins

Comprehensive Overview of Bottom-Up Proteomics Using Mass Spectrometry

The Use of Yeast in Biosensing

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