Easily reversible desthiobiotin binding to streptavidin, avidin, and other biotin-binding proteins: uses for protein labeling, detection, and isolation.
AuthorsHirsch JD, Eslamizar L, Filanoski BJ, Malekzadeh N, Haugland RP, Beechem JM, Haugland RP
JournalAnal Biochem
PubMed ID12419349
'The high-affinity binding of biotin to avidin, streptavidin, and related proteins has been exploited for decades. However, a disadvantage of the biotin/biotin-binding protein interaction is that it is essentially irreversible under physiological conditions. Desthiobiotin is a biotin analogue that binds less tightly to biotin-binding proteins and is easily displaced by ... More
Immunochemical analysis shows all three domains of diphtheria toxin penetrate across model membranes.
AuthorsTortorella D, Sesardic D, Dawes CS, London E
JournalJ Biol Chem
PubMed ID7499201
'Diphtheria toxin undergoes membrane insertion and translocation across membranes when exposed to low pH. In this study, the translocation of the toxin has been investigated by the binding of antibodies to two preparations of model membrane-inserted toxin. In one preparation, toxin was added externally to model membrane vesicles and then ... More
Cell-adhesive properties of streptavidin are mediated by the exposure of an RGD-like RYD site.
AuthorsAlon R, Bayer EA, Wilchek M
JournalEur J Cell Biol
PubMed ID1425765
'The interaction of streptavidin with various cell systems was studied using fluorescent derivatives of the protein. The native unprocessed form of streptavidin bound to cells at low levels and in a nonspecific manner. In contrast, both the truncated "core" streptavidin (the commercially available form) and the biotin-blocked unprocessed protein bound ... More
Cell adhesion to streptavidin via RGD-dependent integrins.
AuthorsAlon R, Bayer EA, Wilchek M
JournalEur J Cell Biol
PubMed ID8462588
'Biotin-blocked streptavidin binds specifically (Kd approximately 3 x 10(-8) M) to cell surfaces, presumably via an RYD-containing sequence. This site is distinct from the biotin-binding cleft of the protein and bears high homology to the RGD-containing cell-binding domain of fibronectin. We show here that various cell types adhere to immobilized ... More
A fluorescence resonance energy transfer-based approach for investigating late endosome-lysosome retrograde fusion events.
AuthorsKaufmann AM, Goldman SD, Krise JP,
JournalAnal Biochem
PubMed ID19109922
'Traditionally, lysosomes have been considered to be a terminal endocytic compartment. Recent studies suggest that lysosomes are quite dynamic, being able to fuse with other late endocytic compartments as well as with the plasma membrane. Here we describe a quantitative fluorescence energy transfer (FRET)-based method for assessing rates of retrograde ... More
Streptavidin contains an RYD sequence which mimics the RGD receptor domain of fibronectin.
AuthorsAlon R, Bayer EA, Wilchek M
JournalBiochem Biophys Res Commun
PubMed ID2390089
'Streptavidin binds at low levels and high affinity to cell surfaces, the cause of which can be traced to the occurrence of a sequence containing RYD (Arg-Tyr-Asp) in the protein molecule. This binding is enhanced in the presence of biotin. Cell-bound streptavidin can be displaced by fibronectin, as well as ... More
Purification and characterization of membrane proteins.
AuthorsKraehenbuhl JP, Bonnard C
JournalMethods Enzymol
PubMed ID2167430
Noncooperativity of biotin binding to tetrameric streptavidin.
AuthorsJones ML, Kurzban GP
JournalBiochemistry
PubMed ID7547907
'Streptavidin tetramers have been separated according to their biotin content by anion exchange chromatography. Biotin-free and biotin-saturated streptavidin were coincubated. Streptavidin at intermediate ligation levels, i.e., with one, two, or three molecules of bound biotin, accumulates over time. A steady state distribution of ligation levels is reached after 2 days. ... More
Fractionation of human lymphocyte subpopulations on immunoglobulin coated Petri dishes.
AuthorsFong S, Tsoukas CD, Pasquali JL, Fox RI, Rose JE, Raiklen D, Carson DA, Vaughan JH
JournalJ Immunol Methods
PubMed ID6456310
'This report describes a simple solid-phase technique for the positive selection of lymphocytes labeled with fluoresceinated antibodies. B lymphocytes were labeled with fluoresceinated anti-human Ig or monoclonal anti-human Ia (L243), and then were bound to plastic culture dishes coated with affinity purified goat anti-fluorescein specific antibody. Bound cells were eluted ... More
A simplified lysis method allowing the use of biotinylated probes in colony hybridization.
AuthorsHaas MJ, Fleming DJ
JournalAnal Biochem
PubMed ID3284412
'A method is described for the lysis of bacterial cells grown on nitrocellulose filters which allows the use of nonradioactive (biotin labeled) probes in colony hybridization. Used in conjunction with a colorimetric assay involving streptavidin and alkaline phosphatase this lysis method allows the detection of clones containing a target nucleic ... More
The use of the avidin-biotin complex as a tool in molecular biology.
AuthorsBayer EA, Wilchek M
JournalMethods Biochem Anal
PubMed ID7392958
Prevention of nonspecific binding of avidin.
AuthorsDuhamel RC, Whitehead JS
JournalMethods Enzymol
PubMed ID2388571
COMBINABILITY OF AVIDIN AND STREPTAVIDIN WITH ANALOGS OF BIOTIN.
AuthorsLICHSTEIN HC, BIRNBAUM J
JournalBiochem Biophys Res Commun
PubMed ID14341938
Detection of low-density cell-surface molecules using biotinylated fluorescent microspheres.
AuthorsWojchowski DM, Sytkowski AJ
JournalBiochim Biophys Acta
PubMed ID3964705
Biotinylated fluorescent microspheres have been developed as a reagent for studying antigens and receptors expressed at the cell surface. Labeling of antigen or receptor was accomplished by crosslinking biotinylated microspheres through streptavidin to corresponding biotinylated antibodies or ligands. Detection of labeled cells by flow microfluorimetry provided an extremely sensitive means ... More
A sensitive enzyme assay for biotin, avidin, and streptavidin.
AuthorsBayer EA, Ben-Hur H, Wilchek M
JournalAnal Biochem
PubMed ID3706735
Reciprocal enzyme assays are described for the vitamin biotin and for the biotin-binding proteins avidin and streptavidin. The assays are based on the following steps: (a) biotinylated bovine serum albumin is adsorbed onto microtiter plates; (b) streptavidin (or avidin) is bound to the biotin-coated plates; (c) biotinylated enzyme (in this ... More
Dissociation rate constant of the biotin-streptavidin complex.
AuthorsPiran U, Riordan WJ
JournalJ Immunol Methods
PubMed ID2212686
We measured the dissociation rate constants of the biotin/streptavidin and biotin/egg avidin complexes by following the release of radiolabeled biotin from the preformed complexes in the presence of excess unlabeled biotin. For separation of bound and free labeled biotin we employed ultrafiltration with disposable microconcentrators. The dissociation rate constant for ... More
Evaluation of biotin-dye conjugates for use in an HPLC assay to assess relative binding of biotin derivatives with avidin and streptavidin.
In this investigation, studies were conducted to determine if size exclusion HPLC could be used to assess relative association rates (on-rates) and dissociation rates (off-rates) of biotin derivatives from avidin (Av) and streptavidin (SAv). For easy detection and quantification of biotin derivatives, molecules that can be detected by UV absorbance ... More
RNA internal standard synthesis by nucleic acid sequence-based amplification for competitive quantitative amplification reactions.
AuthorsLo WY, Baeumner AJ
JournalAnal Chem
PubMed ID17297954
Nucleic acid sequence-based amplification (NASBA) reactions have been demonstrated to successfully synthesize new sequences based on deletion and insertion reactions. Two RNA internal standards were synthesized for use in competitive amplification reactions in which quantitative analysis can be achieved by coamplifying the internal standard with the wild type sample. The ... More