Avidin binding of carboxyl-substituted biotin and analogs

Hofmann, Klaus; Titus, Gail; Montibeller, Judith; Finn, Frances M.

doi:10.1021/bi00534a024

Cited by 138 publications

(88 citation statements)

References 20 publications

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“…Recombinant human HBP was produced using the baculovirus expression system in Sf9 insect cells (Invitrogen Corp., Carlsbad, California, USA) and was purified as described (10). Biotinylation and coupling of HBP to FITC were done as described (17,18). Iodination of HBP was done with N-succinimidyl 3-(4-hydroxy-[ 125 I]iodophenyl)propionate (Amersham International, Amersham, United Kingdom) (19).…”

Section: Methodsmentioning

confidence: 99%

Heparin-binding protein targeted to mitochondrial compartments protects endothelial cells from apoptosis

Olofsson¹,

Vestberg²,

Herwald³

et al. 1999

J. Clin. Invest.

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Section: Methodsmentioning

confidence: 99%

Heparin-binding protein targeted to mitochondrial compartments protects endothelial cells from apoptosis

Olofsson¹,

Vestberg²,

Herwald³

et al. 1999

J. Clin. Invest.

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“…(iii) Biotinylated CAA-specific (2A9) monoclonal antibody (0.2 og/ml PT)was added and incubated for 1 h at 37 °C. This conjugate was prepared by the method of Hofmann et al (1982) using the immunoglobulin fraction of ascites fluid and aminohexanoyl biotin succinate (Zymed). (iv) Streptavidin-peroxidase (Amersham) which had been diluted (1/300) in PT buffer was added and incubated for 30 rain at room temperature.…”

Section: Methodsmentioning

confidence: 99%

Purification and biochemical characterization of chicken anaemia agent

Todd

Creelan

Mackie

et al. 1990

Journal of General Virology

137

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“…To discover such derivatives, we synthesized a series of biotinyl-X insulins (7,21), in which the length of the spacer X was systematically increased, and we measured the rate of dissociation of their complexes with succinoylavidin. The t112 for dissociation of the biotin succinoylavidin complex is 127 days (20).…”

Section: I-labeled Receptor Is Shown Inmentioning

confidence: 99%

Avidin-biotin affinity chromatography: application to the isolation of human placental insulin receptor.

Finn¹,

Titus²,

Horstman³

et al. 1984

Proc. Natl. Acad. Sci. U.S.A.

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The ligand NB'-(6-biotinylamido)hexanoylinsulin was attached noncovalently to Sepharose 4B immobilized succinoylavidin to form an insulin-affinity resin. This resin was used to isolate highly purified insulin receptor from human placental tissue by a four step process involving (i) preparation of a crude membrane fraction, (ii) solubilization with Triton X-100, (iii) wheat germ agglutinin purification, and (iv) insulin-affinity chromatography. NaDodSO4/PAGE of the purified '2I-labeled receptor under nonreducing conditions showed the presence of a major component with an approximate molecular weight of 350,000 and a minor component with a molecular weight of =166,000. Based on the assumption that the degree of labeling is comparable in both components, the material corresponding to the Mr 350,000 peak represents =94% of the receptor preparation as determined by scanning the autoradiograms. The specific insulin binding capacity of the preparation is 18 ± 6 pbg of 12'I-labeled insulin per mg of protein as determined by the polyethylene glycol assay and analyzed by Scatchard plot. Insulin binding activity was stable at 40C and pH 7.6 for at least 12 weeks but was destroyed by freezing and thawing. The availability of highly purified receptor afforded the opportunity to explore its precipitability by polyethylene glycol under assay conditions. Whereas trichloroacetic acid precipitated 95% of the '5I-labeled receptor, polyethylene glycol precipitated only 30%. If the specific activity of the receptor is corrected for incomplete precipitability by polyethylene glycol, the apparent specific binding would be 3.5 + 1.2 mol of insulin per mol of receptor. These results are in disagreement with the current receptor model, which postulates that 1 mol of receptor (Mr, 350,000) binds 2 mol of insulin. Clearly, the problems associated with the method available for determining insulin binding are sufficiently serious to preclude their use in determining receptor valence.Affinity chromatography for insulin receptor isolation was introduced by Cuatrecasas and his colleagues in 1972 (1, 2). An affinity resin was prepared by attaching unprotected insulin to Sepharose via a spacer arm. Using this resin, Jacobs et al. (3) were able to isolate a crude insulin receptor preparation from Triton X-100 solubilized rat liver membranes. The same affinity resin was used by Siegel et al. (4) for the retrieval of insulin receptor preparations from human placenta. The receptor was eluted from the affinity resin with 4.5 M urea (pH 6.0), conditions that damage the receptor. Recently, Fujita-Yamaguchi et al. (5) observed that insulin receptors with high specific insulin binding activity can be eluted from Cuatrecasas-type columns with acetate buffer, pH 5.0/1 M sodium chloride. A considerable store of information concerning the nature of insulin receptors has been accumulated by indirect methods using tracer amounts of materials (6), but methodology for isolating large quantities of receptor is not available. Finite information regarding t...

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Avidin binding of carboxyl-substituted biotin and analogs

Cited by 138 publications

References 20 publications

Heparin-binding protein targeted to mitochondrial compartments protects endothelial cells from apoptosis

Heparin-binding protein targeted to mitochondrial compartments protects endothelial cells from apoptosis

Purification and biochemical characterization of chicken anaemia agent

Avidin-biotin affinity chromatography: application to the isolation of human placental insulin receptor.

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