Ubiquitination and proteasomal activity is required for transport of the EGF receptor to inner membranes of multivesicular bodies.
AuthorsLongva KE, Blystad FD, Stang E, Larsen AM, Johannessen LE, Madshus IH
JournalJ Cell Biol
PubMed ID11864992
'EGF, but not TGF alpha, efficiently induces degradation of the EGF receptor (EGFR). We show that EGFR was initially polyubiquitinated to the same extent upon incubation with EGF and TGF alpha, whereas the ubiquitination was more sustained by incubation with EGF than with TGF alpha. Consistently, the ubiquitin ligase c-Cbl ... More
Optimization of cell surface binding enhances efficiency and specificity of molecular conjugate gene delivery.
AuthorsSchaffer DV, Lauffenburger DA
JournalJ Biol Chem
PubMed ID9774415
'Molecular conjugates, or polyplexes, are promising synthetic vectors for targeted, in vivo gene delivery, if their efficiency can be improved. Gaining mechanistic information on conjugate gene delivery can potentially yield significant improvements in transfer efficiency by revealing barriers to conjugate transfer from the cell surface to the nucleus. We have ... More
A microscale electrospray interface incorporating a monolithic, poly(styrene-divinylbenzene) support for on-line liquid chromatography/tandem mass spectrometry analysis of peptides and proteins.
AuthorsMoore RE, Licklider L, Schumann D, Lee TD
JournalAnal Chem
PubMed ID9852776
'A methodology is described for creating a monolithic chromatography support within a pulled fused-silica electrospray needle. The monolith was formed from a mixture of styrene, divinylbenzene, 1-dodecanol, and toluene using 2,2''-azobis(isobutyronitrile) as the catalyst. The mixture was loaded into 150-micron-i.d. fused-silica capillary tubing with a pulled 5-10-micron needle tip at ... More
Quenching of biotinylated aequorin bioluminescence by dye-labeled avidin conjugates: application to homogeneous bioluminescence resonance energy transfer assays.
AuthorsAdamczyk M, Moore JA, Shreder K
JournalOrg Lett
PubMed ID11405714
[see reaction]. Avidin conjugates containing the covalently attached dyes QSY-7 and dabcyl were prepared and shown to quench the bioluminescence of biotinylated aequorin. Quenching efficiency was shown to be dependent on both the label-to-avidin ratio and the concentration of the avidin conjugate. These properties were exploited to develop a homogeneous ... More
A self-assembled quantum dot probe for detecting beta-lactamase activity.
AuthorsXu C, Xing B, Rao J
JournalBiochem Biophys Res Commun
PubMed ID16631595
This communication describes a quantum dot probe that can be activated by a reporter enzyme, beta-lactamase. Our design is based on the principle of fluorescence resonance energy transfer (FRET). A biotinylated beta-lactamase substrate was labeled with a carbocyanine dye, Cy5, and immobilized on the surface of quantum dots through the ... More
Captavidin: a new regenerable biocomponent for biosensing?
AuthorsGarcía-Aljaro C, Muñoz FX, Baldrich E,
JournalAnalyst
PubMed ID19838424
Functionalisation of a biosensor's sensing surface with the appropriate biorecognition elements is essential for the correct performance of the biosensor. In this paper, we investigate by Surface Plasmon Resonance (SPR) if captavidin, a recently described biotin-binding regenerable protein, could be used to bind and release biotinylated biocomponents for the development ... More
Biotinylation reagents for the study of cell surface proteins.
AuthorsElia G,
JournalProteomics
PubMed ID18763706
The extraordinarily stable, non-covalent interaction between avidin and biotin is one of the most commonly exploited tools in chemistry and biology. Methods for derivatization with biotin of a variety of molecules (in particular, proteins) have been introduced, in order to allow their efficient recovery, immobilization and detection with avidin-based reagents. ... More
Regulation of human CLC-3 channels by multifunctional Ca2+/calmodulin-dependent protein kinase.
AuthorsHuang P, Liu J, Di A, Robinson NC, Musch MW, Kaetzel MA, Nelson DJ
JournalJ Biol Chem
PubMed ID11274166
The multifunctional calcium/calmodulin-dependent protein kinase II, CaMKII, has been shown to regulate chloride movement and cellular function in both excitable and non-excitable cells. We show that the plasma membrane expression of a member of the ClC family of Cl(-) channels, human CLC-3 (hCLC-3), a 90-kDa protein, is regulated by CaMKII. ... More
AP-2/Eps15 interaction is required for receptor-mediated endocytosis.
AuthorsBenmerah A, Lamaze C, Bègue B, Schmid SL, Dautry-Varsat A, Cerf-Bensussan N
JournalJ Cell Biol
PubMed ID9490719
We have previously shown that the protein Eps15 is constitutively associated with the plasma membrane adaptor complex, AP-2, suggesting its possible role in endocytosis. To explore the role of Eps15 and the function of AP-2/Eps15 association in endocytosis, the Eps15 binding domain for AP-2 was precisely delineated. The entire COOH-terminal ... More
Carbachol stimulates binding of a photoreactive calmodulin derivative to calmodulin-binding proteins in intact SK-N-SH human neuroblastoma cells.
AuthorsMangels LA, Gnegy ME
JournalJ Biol Chem
PubMed ID1556101
Calmodulin (CaM) mediates the Ca(2+)-dependent activation of many enzyme systems in accordance with its cellular localization. We have described previously a muscarinic receptor-mediated translocation of CaM from membranes into the cytosol of SK-N-SH human neuroblastoma cells. To explore the potential targets (CaM-binding proteins, CaMBP) for CaM upon translocation, a photoreactive ... More
Molecular and functional characterization of the non-isopeptide-selective ETB receptor in endothelial cells. Receptor coupling to nitric oxide synthase.
AuthorsTsukahara H, Ende H, Magazine HI, Bahou WF, Goligorsky MS
JournalJ Biol Chem
PubMed ID7520443
There is accumulating evidence that endothelial cells express a non-isopeptide-selective endothelin (ET) receptor, ETB, which may be responsible for ET-1-induced transient vasorelaxation. The purpose of the present study was to seek direct evidence for ETB receptor expression in human umbilical vein endothelial cells (HUVEC) and to characterize its functional role ... More