An in vivo imaging-based assay for detecting protein interactions over a wide range of binding affinities

2009 ◽  
Vol 395 (2) ◽  
pp. 166-177 ◽  
Author(s):  
A. Nicole Edwards ◽  
Jason D. Fowlkes ◽  
Elizabeth T. Owens ◽  
Robert F. Standaert ◽  
Dale A. Pelletier ◽  
...  
Keyword(s):  
Protein Interactions ◽  
In Vivo Imaging ◽  
Binding Affinities ◽  
Wide Range

Approach in improving potency and selectivity of kinase inhibitors: Allosteric kinase inhibitors

2020 ◽  
Vol 21 ◽  
Author(s):  
Shangfei Wei ◽  
Tianming Zhao ◽  
Jie Wang ◽  
Xin Zhai
Keyword(s):  
Protein Interactions ◽  
Kinase Inhibitors ◽  
Binding Modes ◽  
Allosteric Inhibitors ◽  
Wide Range ◽  
Allosteric Sites ◽  
Protein Functions ◽  
Regulate Protein

: Allostery is an efficient and particular regulatory mechanism to regulate protein functions. Different from conserved orthosteric sites, allosteric sites have distinctive functional mechanism to form the complex regulatory network. In drug discovery, kinase inhibitors targeting the allosteric pockets have received extensive attention for the advantages of high selectivity and low toxicity. The approval of trametinib as the first allosteric inhibitor validated that allosteric inhibitors could be used as effective therapeutic drugs for treatment of diseases. To date, a wide range of allosteric inhibitors have been identified. In this perspective, we outline different binding modes and potential advantages of allosteric inhibitors. In the meantime, the research processes of typical and novel allosteric inhibitors are described briefly in terms of structureactivity relationships, ligand-protein interactions and in vitro and in vivo activity. Additionally, challenges as well as opportunities are presented.

scholarly journals Ways into Understanding HIF Inhibition

Cancers ◽  
2021 ◽  
Vol 13 (1) ◽  
pp. 159
Author(s):  
Tina Schönberger ◽  
Joachim Fandrey ◽  
Katrin Prost-Fingerle
Keyword(s):  
Protein Interactions ◽  
Target Genes ◽  
Protein Protein Interactions ◽  
Low Oxygen ◽  
Drug Candidates ◽  
Open Questions ◽  
Wide Range ◽  
Hif Pathway

Hypoxia is a key characteristic of tumor tissue. Cancer cells adapt to low oxygen by activating hypoxia-inducible factors (HIFs), ensuring their survival and continued growth despite this hostile environment. Therefore, the inhibition of HIFs and their target genes is a promising and emerging field of cancer research. Several drug candidates target protein–protein interactions or transcription mechanisms of the HIF pathway in order to interfere with activation of this pathway, which is deregulated in a wide range of solid and liquid cancers. Although some inhibitors are already in clinical trials, open questions remain with respect to their modes of action. New imaging technologies using luminescent and fluorescent methods or nanobodies to complement widely used approaches such as chromatin immunoprecipitation may help to answer some of these questions. In this review, we aim to summarize current inhibitor classes targeting the HIF pathway and to provide an overview of in vitro and in vivo techniques that could improve the understanding of inhibitor mechanisms. Unravelling the distinct principles regarding how inhibitors work is an indispensable step for efficient clinical applications and safety of anticancer compounds.

scholarly journals Cofactor-mediated amyloidogenesis

2019 ◽  
Vol 39 (3) ◽  
Author(s):  
Jillian Madine
Keyword(s):  
Conformational Change ◽  
Conformational Changes ◽  
Protein Interactions ◽  
Zinc Binding ◽  
Amyloid Protein ◽  
Wide Range ◽  
Pathogenesis Related Protein ◽  
Plant Pathogenesis

Abstract A recent study published in Bioscience Reports by Sheng et al. (Bioscience Reports, (2019) 39, pii:BSR20182345] described a small but significant conformational change that occurs upon zinc binding and results in initiation of the amyloidogenic aggregation cascade of Golgi-Associated plant Pathogenesis Related protein 1 (GAPR-1) in the presence of heparin. The present study describes a two-stage process that is required for the initiation of the amyloidogenic aggregation cascade involving a concentration step and a conformation change to enhance accessibility of natively protected amyloidogenic regions for self-association. For GAPR-1 in the present study, these steps are provided by zinc binding causing the required conformational change enhancing accessibility of amyloidogenic regions, and heparin providing a template or scaffold in turn increasing the local protein concentration. Cofactors such as glycosaminoglycans and metal ions have been found associated with amyloid deposits in vivo and shown to affect protein assembly kinetics in vitro. Cofactor interactions with the amyloidogenic process are an area of great interest for therapeutic intervention for the wide range of diseases known to be associated with amyloid protein aggregation. The present study emphasises the need for enhanced structural understanding of cofactor–amyloid protein interactions and highlights that small subtle conformational changes can have large impacts on resulting aggregation processes.

Nanotechnology: Better Materials for All Implants

2007 ◽  
Vol 539-543 ◽  
pp. 511-516 ◽  
Author(s):  
Thomas J. Webster
Keyword(s):  
Protein Interactions ◽  
Review Paper ◽  
Short Review ◽  
Structural Components ◽  
Synthetic Materials ◽  
Wide Range ◽  
Mediate Cell Adhesion ◽  
Mediate Cell

Nanotechnology is being used to mimic structural components of our tissues in synthetic materials intended for various implant applications. Recent studies have highlighted that when compared to flat or micron rough surfaces, surfaces with nanofeatures promote optimal initial protein interactions necessary to mediate cell adhesion and subsequent tissue regrowth. This has been demonstrated for a wide range of implant chemistries (from ceramics to metals to polymers) and for a wide range of tissues (including bone, vascular, cartilage, bladder, and the central and peripheral nervous system). Importantly, these results have been seen at the in vitro and in vivo level. This short review paper will cover some of the more significant advancements in creating better implants through nanotechnology efforts.

scholarly journals Global query on bacterial sirtuin CobB interactants reveals crosstalk with PRPP synthase Prs

2020 ◽  
Author(s):  
Beata M. Walter ◽  
Joanna Morcinek-Orłowska ◽  
Aneta Szulc ◽  
Andrew L. Lovering ◽  
Manuel Banzhaf ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Global Analysis ◽  
Stable Complex ◽  
Protein Protein Interactions ◽  
Cellular Functions ◽  
Nad Metabolism ◽  
E Coli ◽  
Nad Synthesis ◽  
Wide Range

AbstractProtein lysine acetylation regulates a wide range of cellular functions. It is controlled by a family of NAD-dependent protein deacetylases called sirtuins. In eukaryotes, sirtuins activity is coupled to spatiotemporally-controlled NAD+ level, whereas the mechanism of their regulation in bacteria is less clear. E. coli possesses a single sirtuin – CobB. However, it is unclear how CobB activity is coupled to NAD+ metabolism. In this work we show that this coordination is achieved in E. coli cells through a CobB interaction with PRPP synthase Prs, an enzyme necessary for NAD+ synthesis. Employing global analysis of protein-protein interactions formed by CobB, we demonstrate that it forms a stable complex with Prs. This assembly stimulates CobB deacetylase activity and partially protects it from inhibition by nicotinamide. We provide evidence that Prs acetylation is not necessary for CobB binding but affects the global acetylome in vivo. Our results show that CobB ameliorates Prs activity under conditions of Prs cofactors deficiency. Therefore, we propose that CobB-Prs crosstalk orchestrates the NAD+ metabolism and protein acetylation in response to environmental cues.

scholarly journals A multi-reporter bacterial 2-hybrid assay for the fast, simple, and dynamic assay of PDZ domain – peptide interactions

2019 ◽  
Author(s):  
David M. Ichikawa ◽  
Carles Corbi-Verge ◽  
Michael J. Shen ◽  
Jamie Snider ◽  
Victoria Wong ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Accurate Determination ◽  
Pdz Domain ◽  
Relative Strength ◽  
Protein Protein Interactions ◽  
Reporter System ◽  
Protein Motif ◽  
Wide Range ◽  
Domain Peptide

AbstractThe accurate determination of protein-protein interactions is an important goal of molecular biology and much progress has been made with modern methods emerging in the past decade. However, current methods have limitations, including scale and restriction to high affinity interactions. This has limited our understanding of the large subset of protein-motif interactions. Here we describe a modified bacterial-hybrid assay that employs a combined selectable and scalable reporter system that allows the screen of large protein-peptide libraries and their sort by relative strength. We have applied this tool to characterize a set of human and E. coli PDZ domains. Our results are consistent with prior characterization of these proteins and the improved sensitivity increases our ability to predict known and novel in vivo binding partners. This approach allows for the recovery of a wide range of affinities with a high throughput method that does not sacrifice the scale of the screen.

scholarly journals Systematic discovery of Short Linear Motifs decodes calcineurin phosphatase signaling

2019 ◽  
Author(s):  
Callie P. Wigington ◽  
Jagoree Roy ◽  
Nikhil P. Damle ◽  
Vikash K. Yadav ◽  
Cecilia Blikstad ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Nuclear Pore ◽  
Binding Affinities ◽  
Protein Protein Interactions ◽  
Short Linear Motifs ◽  
Binding Peptides ◽  
Linear Motifs ◽  
Calcineurin Phosphatase

SummaryShort linear motifs (SLiMs) drive dynamic protein-protein interactions essential for signaling, but sequence degeneracy and low binding affinities make them difficult to identify. We harnessed unbiased systematic approaches for SLiM discovery to elucidate the regulatory network of calcineurin (CN)/PP2B, the Ca2+-activated phosphatase that recognizes LxVP and PxIxIT motifs. In vitro proteome-wide detection of CN-binding peptides, in vivo SLiM-dependent proximity labeling, and in silico modeling of motif determinants uncovered unanticipated CN interactors, including NOTCH1, which we establish as a CN substrate. Unexpectedly, CN shows SLiM-dependent proximity to centrosomal and nuclear pore complex (NPC) proteins – structures where Ca2+ signaling is largely uncharacterized. CN dephosphorylates human and yeast NPC proteins and promotes accumulation of a nuclear transport reporter, suggesting conserved NPC regulation by CN. The CN network assembled here provides a resource to investigate Ca2+ and CN signaling and demonstrates synergy between experimental and computational methods, establishing a blueprint for examining SLiM-based networks.

scholarly journals In Vivo Imaging With Confirmation by Histopathology for Increased Rigor and Reproducibility in Translational Research: A Review of Examples, Options, and Resources

ILAR Journal ◽  
2018 ◽  
Vol 59 (1) ◽  
pp. 80-98 ◽  
Author(s):  
Kathleen Gabrielson ◽  
Robert Maronpot ◽  
Sébastien Monette ◽  
Coraline Mlynarczyk ◽  
Yuval Ramot ◽  
...  
Keyword(s):  
Animal Model ◽  
In Vivo Imaging ◽  
Image Interpretation ◽  
Imaging Data ◽  
Case Examples ◽  
Animal Models Of Disease ◽  
Molecular Features ◽  
Wide Range ◽  
Models Of Disease

Abstract Preclinical noninvasive imaging can be an indispensable tool for studying animal models of disease. In vivo imaging to assess anatomical, functional, and molecular features requires verification by a comparison to the macroscopic and microscopic morphological features, since all noninvasive in vivo imaging methods have much lower resolution than standard histopathology. Comprehensive pathological evaluation of the animal model is underutilized; yet, many institutions have veterinary or human pathologists with necessary comparative pathology expertise. By performing a rigorous comparison to gross or histopathology for image interpretation, these trained individuals can assist scientists with the development of the animal model, experimental design, and evaluation of the in vivo imaging data. These imaging and pathology corroboration studies undoubtedly increase scientific rigor and reproducibility in descriptive and hypothesis-driven research. A review of case examples including ultrasound, nuclear, optical, and MRI is provided to illustrate how a wide range of imaging modalities data can be confirmed by gross or microscopic pathology. This image confirmation and authentication will improve characterization of the model and may contribute to decreasing costs and number of animals used and to more rapid translation from preclinical animal model to the clinic.

Sign in / Sign up

Export Citation Format

Share Document