scholarly journals A novel mechanism-based mammalian cell assay for the identification of SH2-domain-specific protein-protein inhibitors

1998 ◽  
Vol 5 (10) ◽  
pp. 529-538 ◽  
Author(s):  
Richard J. Rickles ◽  
Pamela A. Henry ◽  
Wei Guan ◽  
Mihai Azimioara ◽  
William C. Shakespeare ◽  
...  
Keyword(s):  
Mammalian Cell ◽  
Specific Protein ◽  
Sh2 Domain ◽  
Protein Inhibitors ◽  
Domain Specific ◽  
Cell Assay

scholarly journals Novel Roles of SH2 and SH3 Domains in Lipid Binding

Cells ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 1191
Author(s):  
Szabolcs Sipeki ◽  
Kitti Koprivanacz ◽  
Tamás Takács ◽  
Anita Kurilla ◽  
Loretta László ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Specific Protein ◽  
Lipid Binding ◽  
Sh2 Domain ◽  
Essential Property ◽  
Cellular Regulation ◽  
Sh3 Domains ◽  
Protein Partners ◽  
Interaction Domains ◽  
Central Paradigm

Signal transduction, the ability of cells to perceive information from the surroundings and alter behavior in response, is an essential property of life. Studies on tyrosine kinase action fundamentally changed our concept of cellular regulation. The induced assembly of subcellular hubs via the recognition of local protein or lipid modifications by modular protein interactions is now a central paradigm in signaling. Such molecular interactions are mediated by specific protein interaction domains. The first such domain identified was the SH2 domain, which was postulated to be a reader capable of finding and binding protein partners displaying phosphorylated tyrosine side chains. The SH3 domain was found to be involved in the formation of stable protein sub-complexes by constitutively attaching to proline-rich surfaces on its binding partners. The SH2 and SH3 domains have thus served as the prototypes for a diverse collection of interaction domains that recognize not only proteins but also lipids, nucleic acids, and small molecules. It has also been found that particular SH2 and SH3 domains themselves might also bind to and rely on lipids to modulate complex assembly. Some lipid-binding properties of SH2 and SH3 domains are reviewed here.

scholarly journals Development of Binding Assays for the SH2 Domain of Grb7 and Grb2 Using Fluorescence Polarization

2008 ◽  
Vol 13 (2) ◽  
pp. 112-119 ◽  
Author(s):  
Jean-Philippe Luzy ◽  
Huixiong Chen ◽  
Brunilde Gril ◽  
Wang-Qing Liu ◽  
Michel Vidal ◽  
...  
Keyword(s):  
Fluorescence Polarization ◽  
Adaptor Proteins ◽  
Specific Protein ◽  
Sh2 Domain ◽  
Enzyme Linked Immunosorbent Assay ◽  
Binding Assays ◽  
Protein Targets ◽  
Src Homology 2 ◽  
Src Homology ◽  
Micromolar Range

Adaptor proteins Grb7 and Grb2 have been implicated as being 2 potential therapeutic targets in several human cancers, especially those that overexpress ErbB2. These 2 proteins contain both a SH2 domain (Src homology 2) that binds to phosphorylated tyrosine residues contained within ErbB2 and other specific protein targets. Two assays based on enzyme-linked immunosorbent assay and fluorescence polarization methods have been developed and validated to find and rank inhibitors for both proteins binding to the pY1139. Fluorescence polarization assays allowed the authors to determine quickly and reproducibly affinities of peptides from low nanomolar to high micromolar range and to compare them directly for Grb7 and Grb2. As a result, the assays have identified a known peptidomimetic Grb2 SH2 inhibitor (mAZ-pTyr-(αMe)pTyr-Asn-NH2) that exhibits the most potent affinity for the Grb7 SH2 domain described to date. ( Journal of Biomolecular Screening 2008:112-119)

Genotoxicity of fecapentaene-12 in bacterial and mammalian cell assay systems

1987 ◽  
Vol 8 (2) ◽  
pp. 349-352 ◽  
Author(s):  
R.D. Curren ◽  
D.L Putman ◽  
L.L Yang ◽  
S.R Haworth ◽  
T.E Lawlor ◽  
...  
Keyword(s):  
Mammalian Cell ◽  
Cell Assay

scholarly journals The SH2 domain of P210BCR/ABL is not required for the transformation of hematopoietic factor-dependent cells

Blood ◽  
1995 ◽  
Vol 86 (10) ◽  
pp. 3897-3904 ◽  
Author(s):  
RL Jr Ilaria ◽  
RA Van Etten
Keyword(s):  
Cell Lines ◽  
Protein Interactions ◽  
Specific Protein ◽  
Sh2 Domain ◽  
Myelogenous Leukemia ◽  
Mutant Proteins ◽  
Abl Tyrosine Kinase ◽  
Negative Effect ◽  
Dependent Cell

Src-homology region 2 (SH2) domains, by binding to tyrosine- phosphorylated sequences, mediate specific protein-protein interactions important in diverse signal transduction pathways. Previous studies have shown that activated forms of the Abl tyrosine kinase, including P210BCR/ABL of human chronic myelogenous leukemia, require the SH2 domain for the transformation of fibroblasts. To determine whether SH2 is also required for Bcr/Abl to transform hematopoietic cells, we have studied two SH2 domain mutations in P210BCR/ABL: a point mutation in the conserved FLVRES motif (P210/R1033K), which interferes with phosphotyrosine-binding by SH2, and a complete deletion of SH2 (P210/delta SH2). Despite a negative effect on intrinsic Abl kinase activity, both P210 SH2 mutants were still able to transform the hematopoietic factor-dependent cell lines Ba/F3 and FDC-P1 to growth factor independence. Unexpectedly, both mutants showed greater transforming activity than wild-type P210 in a quantitative transformation assay, probably as a consequence of increased stability of the SH2 mutant proteins in vivo. Cells transformed by both P210 SH2 mutants were leukemogenic in synaptic mice and P210/r1053K mice exhibited a distinct disease phenotype, reminiscent of that induced by v-Abl. These results demonstrate that while the Abl SH2 domain is essential for BCR/ABL transformation of fibroblasts, it is dispensable for the transformation of hematopoietic factor-dependent cell lines.

scholarly journals Cyclodextrin polymer coatings resist protein fouling, mammalian cell adhesion, and bacterial attachment

2020 ◽  
Author(s):  
Greg D. Learn ◽  
Emerson J. Lai ◽  
Horst A. von Recum
Keyword(s):  
Mammalian Cell ◽  
Pharmaceutical Formulations ◽  
Polymer Coatings ◽  
Specific Protein ◽  
Bacterial Attachment ◽  
Polymer Composition ◽  
Passive Resistance ◽  
Cyclodextrin Polymer ◽  
Sustained Drug Delivery ◽  
Nih 3T3

AbstractUndesired attachment of proteins, cells/bacteria, and organisms on material surfaces is problematic in industrial and health care settings. In this study, polymer coatings are synthesized from subunits of cyclodextrin, an additive/excipient found in food/pharmaceutical formulations. These unique polymers, which have been applied mainly towards sustained drug delivery applications, are evaluated in this study for their ability to mitigate non-specific protein adsorption, mammalian cell (NIH/3T3) adhesion, and bacterial cell (Staphylococcus aureus, Escherichia coli) attachment. Effects of cyclodextrin polymer composition, particularly incorporation of nonpolar crosslinks, on material properties and passive anti-biofouling performance are investigated. Results suggest that lightly-crosslinked cyclodextrin polymers possess excellent passive resistance to protein, cell, and bacterial attachment, likely due to the hydrophilic and electrically neutral surface properties of these coatings. At the same time, anti-biofouling performance decreased with increasing crosslink ratios, possibly a reflection of decreased polymer mobility, increased rigidity, and increased hydrophobic character. Cyclodextrin-based materials may be broadly useful as coatings in industrial or medical applications where biofouling-resistant and/or drug-delivering surfaces are required.

Effects of Proteinase Inhibitors and Plant Lectins on the Adult Alfalfa Weevil (Coleoptera: Curculionidae)

2000 ◽  
Vol 35 (1) ◽  
pp. 62-69 ◽  
Author(s):  
T. C. Elden
Keyword(s):  
Adult Stage ◽  
Proteinase Inhibitors ◽  
Cysteine Proteinase ◽  
Specific Protein ◽  
Adult Survival ◽  
Protein Inhibitors ◽  
Alfalfa Weevil ◽  
Hypera Postica ◽  
Plant Lectins ◽  
Cysteine Proteinase Inhibitors

The effects of selected proteinase inhibitors and plant lectins of alfalfa weevil, Hypera postica (Gyllenhal), adult foliar feeding and fecundity were significantly inhibited by the cysteine proteinase inhibitors E-64, pHMB, and leupeptin at a concentration of 0.1%. Pepstatin (aspartic inhibitor) at 0.5% and soybean Bowman-Birk trypsin inhibitor (serine) at 1.0% had no significant effect on adult foliar feeding, survival, or fecundity. Three of the four lectins tested significantly inhibited adult foliar feeding and fecundity at a concentration of 0.5%. A lectin from wheat and one from pea were the only two protein inhibitors tested to significantly inhibit adult survival. Results support a previous study that indicates the alfalfa weevil uses cysteine proteinases as major digestive enzymes. This study is one of few which demonstrates the effects of specific protein inhibitors on the adult stage of a foliar feeding insect species.

MAMMALIAN CELL ASSAY SYSTEMS FOR ENDOCRINE DISRUPTING CHEMICALS

2000 ◽  
Vol 28 (5) ◽  
pp. A435-A435
Author(s):  
P. P. Darbre ◽  
J. R. Higman ◽  
L. Shaw ◽  
M. J. Sauer
Keyword(s):  
Mammalian Cell ◽  
Endocrine Disrupting Chemicals ◽  
Cell Assay ◽  
Endocrine Disrupting

scholarly journals Dissecting lipid contents in the distinct regions of native retinal rod disk membranes

2021 ◽  
Author(s):  
Christopher L. Sander ◽  
Avery E. Sears ◽  
Antonio F. M. Pinto ◽  
Elliot H. Choi ◽  
Shirin Kahremany ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Membrane Proteins ◽  
Lipid Composition ◽  
Protein Localization ◽  
Specific Protein ◽  
Fatty Acid Analysis ◽  
Protein Activity ◽  
Domain Specific ◽  
Retinal Rod ◽  
Membrane Compartments

ABSTRACTPhotoreceptors rely on distinct membrane compartments to support their specialized function. Unlike protein localization, identification of critical differences in membrane content has not yet been expanded to lipids, due to the difficulty of isolating domain-specific samples. We have overcome this by using SMA to co-immunopurify membrane proteins and their native lipids from two regions of photoreceptor ROS disks. Each sample’s copurified lipids were subjected to untargeted lipidomic and fatty acid analysis. Extensive differences between center (rhodopsin) and rim (ABCA4 and PRPH2/ROM1) samples included a lower PC to PE ratio and increased LC- and VLC-PUFAs in the center relative to the rim region, which were enriched in shorter, saturated FAs. The comparatively few differences between the two rim samples likely reflect specific protein-lipid interactions. High-resolution profiling of the ROS disk lipid composition provides a model for future studies of other complex cellular structures, and gives new insights into how intricate membrane structure and protein activity are balanced within the ROS.SUMMARYSander et al. have parsed the lipid composition of native-source photoreceptor disks and find large differences in fatty acid unsaturation and chain length between the center and rim regions. They selectively copurify membrane proteins and lipids from each region in SMALPs using nanobodies and antibodies.

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