A versatile method of identifying specific binding proteins on affinity resins

2006 ◽  
Vol 352 (1) ◽  
pp. 15-23 ◽  
Author(s):  
Kiyoshi Yamamoto ◽  
Akira Yamazaki ◽  
Mikio Takeuchi ◽  
Akito Tanaka
Keyword(s):  
Binding Proteins ◽  
Specific Binding

scholarly journals Identification of a fibroblast growth factor-binding protein in Drosophila melanogaster.

1991 ◽  
Vol 11 (4) ◽  
pp. 2319-2323 ◽  
Author(s):  
J S Doctor ◽  
F M Hoffmann ◽  
B B Olwin
Keyword(s):  
Molecular Weight ◽  
Drosophila Melanogaster ◽  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Binding Proteins ◽  
Specific Binding ◽  
Fibroblast Growth ◽  
Heparin Binding ◽  
Acidic Fibroblast Growth Factor ◽  
Heparin Binding Growth Factors

As assessed by competitive binding and protein-crosslinking experiments, Drosophila melanogaster cells possess basic fibroblast growth factor (bFGF)-specific binding proteins that are similar to FGF receptors on vertebrate cells in molecular weight and binding affinity; these D. melanogaster cells, however, have no detectable binding proteins for acidic fibroblast growth factor (aFGF). Consistent with the presence of bFGF-specific binding proteins, D. melanogaster cells degrade bFGF but not aFGF. These results indicate the conservation of heparin-binding growth factors and receptors between vertebrates and D. melanogaster.

The induction of a specific protease for insulin-like growth factor binding protein-3 in the circulation during severe illness

1991 ◽  
Vol 130 (3) ◽  
pp. 469-NP ◽  
Author(s):  
S. C. Davies ◽  
J. A. H. Wass ◽  
R. J. M. Ross ◽  
A. M. Cotterill ◽  
C. R. Buchanan ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Specific Binding ◽  
Nitrogen Retention ◽  
Late Pregnancy ◽  
Severe Illness ◽  
Igf I ◽  
Catabolic State ◽  
Specific Protease ◽  
Circulating Levels ◽  
Igfbp 3

ABSTRACT The insulin-like growth factors (IGF-I and IGF-II) are almost completely bound in the circulation to specific binding proteins (IGFBPs). These IGFBPs appear to play a pivotal role in maintaining circulating levels and modulating the delivery of the IGFs to the tissues. A large proportion of the circulating IGFs are bound with high affinity to one of the binding proteins, IGFBP-3. The mechanism by which these IGFs are transferred from the circulatory pool to the tissue receptors is at present unclear. Recent studies in late pregnancy have demonstrated the presence of specific proteases which may modify the IGFBPs such that their affinities for the IGFs are reduced. In this paper, we have demonstrated the presence of a heat-sensitive cation-dependent proteolytic enzyme specific for IGFBP-3 in the serum of five severely ill patients. The activity of this protease was found to vary in these patients, becoming more apparent during fasting than when studied after commencement of parenteral nutrition, indicating that one of the influencing factors in the activity of this protease is the nutritional intake of the patient. Age- and sex-matched healthy adults were also studied in a similar protocol, but no proteolytic modification of any of the IGFBPs was found in any of the samples examined. As the levels of both IGF-I and IGF-II were found to be low in the patients, the presence of a circulatory protease suggests that this may be an adaptive response to increase the bioavailability of the IGFs and possibly to improve the nitrogen retention and counter the catabolic state in severe illness. Journal of Endocrinology (1991) 130,469–473

Enhancement by GTP of cAMP binding to hepatic nuclei and cytosol

1984 ◽  
Vol 246 (1) ◽  
pp. C131-C140 ◽  
Author(s):  
E. M. Rosenberg ◽  
A. D. Goodman ◽  
T. L. Lipinski
Keyword(s):  
Binding Proteins ◽  
Specific Binding ◽  
Photoaffinity Labeling ◽  
Deae Cellulose ◽  
Nuclear Extract ◽  
Liver Cytosol ◽  
Nuclear Binding ◽  
Low Concentrations ◽  
Relative Molecular Weight

In the present study we have demonstrated specific binding of 3H-labeled adenosine 3',5'-cyclic monophosphate (cAMP) to a nuclear extract from rat liver. GTP, GDP, and low concentrations of ATP and ADP increased nuclear binding of [3H]cAMP, and AMP inhibited [3H]cAMP binding. Photoaffinity labeling studies employing [32P]cAMP revealed four nuclear binding proteins [relative molecular weight (Mr) 36,000, 49,000, 54,000 and 57,000]. Unlabeled cAMP decreased [32P]cAMP binding to all four proteins, whereas GTP increased binding to the 57,000 protein. We also observed specific binding of [3H]cAMP in the liver cytosol, which was stimulated by GTP but not by ADP or ATP. Photoaffinity labeling studies of the cytosol in the absence of unlabeled nucleotides revealed three cAMP-binding proteins (Mr 36,000, 49,000, and 54,000). Unlabeled cAMP inhibited binding of [32P]cAMP to all three proteins, whereas in the presence of GTP there was binding of [32P]cAMP to a Mr 57,000 protein. Using DEAE-cellulose, we isolated from the nuclear extract and cytosol a cAMP-binding protein that responded to GTP with an increase in cAMP binding but was unaffected by GDP, ATP, ADP, and AMP. Guanosine imidodiphosphate did not affect cAMP binding, suggesting that the stimulatory effect of GTP may be mediated by phosphorylation. We speculate that alterations in intracellular GTP in vivo may modulate the binding of cAMP to a protein in the nucleus and cytosol.

scholarly journals Reading RNA methylation codes through methyl-specific binding proteins

RNA Biology ◽  
2014 ◽  
Vol 11 (6) ◽  
pp. 669-672 ◽  
Author(s):  
Xiao Wang ◽  
Chuan He
Keyword(s):  
Binding Proteins ◽  
Specific Binding ◽  
Rna Methylation

scholarly journals GTP-binding proteins may stimulate insulin biosynthesis in rat pancreatic islets by enhancing the signal-recognition-particle-dependent translocation of the insulin mRNA poly-/mono-some complex to the endoplasmic reticulum

1991 ◽  
Vol 275 (1) ◽  
pp. 23-28 ◽  
Author(s):  
N Welsh ◽  
C Oberg ◽  
M Welsh
Keyword(s):  
Endoplasmic Reticulum ◽  
Binding Proteins ◽  
Specific Binding ◽  
Signal Recognition Particle ◽  
Insulin Biosynthesis ◽  
Signal Recognition ◽  
Insulin Synthesis ◽  
Gtp Binding Proteins ◽  
Gtp Binding ◽  
Insulin Mrna

We aimed to elucidate the putative role of GTP-binding proteins in the regulation of insulin biosynthesis. For this purpose, freshly isolated rat islets were incubated in the presence of liposomes containing GDP, guanosine 5′-[beta-thio]diphosphate (GDP[S]), GTP, guanosine 5′-[gamma-thio]triphosphate (GTP[S]), guanosine 5′-[beta gamma-methylene]triphosphate (p[CH2]ppG), guanosine 5′[beta gamma-imido]triphosphate (p[NH]ppG) and ATP, and the effects of the liposomal delivery of these substances on rates of biosynthesis of insulin and total protein were determined. Insulin biosynthesis during a 1 h incubation at 1.67 mM-glucose was stimulated by ATP- and GTP[S]-containing liposomes as compared with control liposomes. At 16.7 mM-glucose, only the GTP[S]-containing liposomes stimulated insulin biosynthesis. No inhibition of islet protein and insulin synthesis was observed with GDP-, GDP[S]-, p[CH2]ppG- and p[NH]ppG-containing liposomes. By determining the subcellular distribution of insulin mRNA, it was found that the mRNA content associated with microsomes was increased and that associated with the cytosolic mono-/poly-somes decreased when the islets were incubated with GTP[S]-containing liposomes, resulting in an approximate doubling of the ratio of microsomal to polysomal-associated insulin mRNA. ATP-containing liposomes produced no effects on the association of insulin mRNA with microsomes. By using photoaffinity labelling and immunoprecipitation techniques, specific binding of GTP[35S] to the alpha-subunit of the signal-recognition particle (SRP) receptor in islet homogenates containing physiological concentrations of GTP and GDP was demonstrated. These findings suggest that the GTP-binding subunit(s) of the SRP receptor, and possibly also of other GTP-binding proteins involved in this process, may regulate insulin biosynthesis by stimulating the translocation of insulin mRNA to the endoplasmic reticulum and by increasing preproinsulin-peptide translocation into the lumen of the reticulum.

Specific binding proteins for selenium in rat tissues

1988 ◽  
Vol 9 (2) ◽  
pp. 277-284 ◽  
Author(s):  
Brahma P. Sani ◽  
John L. Woodard ◽  
Milton C. Pierson ◽  
Roger D. Allen
Keyword(s):  
Binding Proteins ◽  
Specific Binding ◽  
Rat Tissues

scholarly journals Synthesis and application of photoaffinity analogues of inositol 1,4,5-trisphosphate selectively substituted at the 1-phosphate group

1990 ◽  
Vol 272 (3) ◽  
pp. 817-825 ◽  
Author(s):  
R Schäfer ◽  
M Nehls-Sahabandu ◽  
B Grabowsky ◽  
M Dehlinger-Kremer ◽  
I Schulz ◽  
...  
Keyword(s):  
Binding Sites ◽  
Binding Proteins ◽  
Inositol Phosphates ◽  
Specific Binding ◽  
Acinar Cells ◽  
Phosphate Group ◽  
Pancreatic Acinar Cells ◽  
Maximal Effect ◽  
Microsomal Preparation ◽  
Specific Binding Sites

We have synthesized two photolabile arylazido-analogues of Ins(1,4,5)P3 selectively substituted at the 1-phosphate group for determination of Ins(1,4,5)P3-binding proteins. These two photoaffinity derivatives, namely N-(4-azidobenzoyl)aminoethanol-1-phospho-D-myo-inositol 4,5-bisphosphate (AbaIP3) and N-(4-azidosalicyl)aminoethanol-1-phospho-D-myo-inositol 4,5-bisphosphate (AsaIP3), bind to high affinity Ins(1,4,5)P3-specific binding sites at a 9-fold lower affinity (Kd = 66 and 70 nM) than Ins(1,4,5)P3 (Kd = 7.15 nM) in a fraction from rat pancreatic acinar cells enriched in endoplasmic reticulum (ER). Other inositol phosphates tested showed comparable (DL-myo-inositol 1,4,5-trisphosphothioate, Kd = 81 nM) or much lower affinities for the binding sites [Ins(1,3,4,5)P4, Kd = 4 microM; Ins(1,4)P2, Kd = 80 microM]. Binding of AbaIP3 was also tested on a microsomal preparation of rat cerebellum [Kd = 300 nM as compared with Ins(1,4,5)P3, Kd = 45 nM]. Ca2+ release activity of the inositol derivatives was tested with AbaIP3. It induced a rapid and concentration-dependent Ca2+ release from the ER fraction [EC50 (dose producing half-maximal effect) = 3.1 microM] being only 10-fold less potent than Ins(1,4,5)P3 (EC50 = 0.3 microM). From the two radioactive labelled analogues ([3H]AbaIP3 and 125I-AsIP3) synthesized, the radioiodinated derivative was used for photoaffinity labelling. It specifically labelled three proteins with apparent molecular masses of 49, 37 and 31 kDa in the ER-enriched fraction. By subfractionation of this ER-enriched fraction on a Percoll gradient the 37 kDa Ins(1,4,5)P3 binding protein was obtained in a membrane fraction which showed the highest effect in Ins(1,4,5)P3-inducible Ca2+ release (fraction P1). The other two Ins(1,4,5)P3-binding proteins, of 49 and 31 kDa, were obtained in fraction P2, in which Ins(1,4,5)P3-induced Ca2+ release was half of that obtained in fraction P1. We conclude from these data that the 37 kDa and/or the 49 and 31 kDa proteins are involved in Ins(1,4,5)P3-induced Ca2+ release from the ER of rat pancreatic acinar cells.

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