Specific binding of free apolipoprotein A-I to a high-affinity binding site on HepG2 Cells: characterization of two high-density lipoprotein sites

Biochemistry ◽  
1994 ◽  
Vol 33 (8) ◽  
pp. 2335-2340 ◽  
Author(s):  
Ronald Barbaras ◽  
Xavier Collet ◽  
Hugues Chap ◽  
Bertrand Perret
Keyword(s):  
High Density Lipoprotein ◽  
Hepg2 Cells ◽  
Specific Binding ◽  
Density Lipoprotein ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity ◽  
Apolipoprotein A ◽  
High Affinity Binding Site

Opioid receptors in bovine adrenal medulla

1984 ◽  
Vol 62 (10) ◽  
pp. 1284-1291 ◽  
Author(s):  
Michel Dumont ◽  
Simon Lemaire
Keyword(s):  
Binding Site ◽  
G Protein ◽  
Opioid Receptors ◽  
Opioid Peptides ◽  
Affinity Binding ◽  
Endogenous Opioid ◽  
High Affinity Binding ◽  
High Affinity ◽  
High Affinity Binding Site

Using prototypic ligands for each type of opioid receptors (μ, δ, κ, and σ) as well as compounds derived from each class of endogenous opioid peptides (β-endorphin, enkephalins, and dynorphins), we have undertaken the characterization of adrenomedullary opioid binding sites. The specific binding of [3H]etorphine ([3H]ET) to a membrane preparation of bovine adrenal medulla was greatly increased when the incubation temperature was raised from 22 to 37 °C. Characterization of the opioid binding sites was obtained at 37 °C with [3H]ET (nonspecific opioid ligand), [3H]ethylketocyclazocine ([3H]EKC; κ), [3H]dihydromorphine ([3H]DHM; μ), [3H]-[D-Ala2,D-Leu5]enkephalin ([3H]DADLE; δ), and N-[3H]allylnormetazocine ([3H]SKF-10047; σ) in the absence or presence of blocking agents for cross-reacting receptors. [3H]ET had a high affinity binding site (KD = 0.98 nM) with a Bmax of 119 pmol/g protein. All the other opioid compounds showed biphasic saturation curves with KD ranging from 0.6 to 1.29 nM for the high affinity binding site and from 2.49 to 12.1 nM for the low affinity binding site. The opioid μ-receptor was characterized by the high affinity binding site for [3H]DHM (KD = 1.29 nM; Bmax = 38 pmol/g protein). Blockade of the cross-reacting receptor sites for [3H]EKC, [3H]DADLE, and [3H]SKF-10047 revealed the presence of κ (KD = 0.66 nM; Bmax = 12 pmol/g protein), κ2 (benzomorphan site; KD = 11.1 nM; Bmax = 56 pmol/g protein), δ (KD = 0.67 nM; Bmax = 4.7 pmol/g protein), and σ (KD = 4.54 nM; Bmax = 32 pmol/g protein) opioid receptors. The ability of various opioid ligands to displace the binding of [3H]ET indicates a high potency for (−)-(1R,5R,9R,2″S)-5,9-dimethyl-2′-hydroxy-2-tetrahydrofurfuryl-6,7-benzomorphan hydrogen D-tartrate (MR-2034, a κ-opioid ligand; Ki = 6.2 nM), dihydromorphinone (DHMone; Ki = 6.9 nM), oxymorphone (Ki = 8.6 nM), DADLE (Ki high affinity = 8.4 nM) EKC (Ki = 31.8 nM), SKF-10047 (Ki = 75 nM), and opioid agonists/antagonists. trans-(+)-3,4-Dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl]benzeneacetamide methanesulfonate hydrate (U-50,488H), the most specific κ-agonist, was a poor competitor (Ki = 5150 nM). However, the presence of κ-opioid receptors was supported by the ability of U-50,488H to displace [3H]EKC binding (Ki high affinity = 2.5 nM). The relative potency of various endogenous opioid peptides in displacing [3H]ET binding was as follows: β-endorphin [Formula: see text] dynorphin(1-17) > dynorphin(1-13) > [Arg6,Phe7)Met-enkephalin > Met-enkephalin > Leu-enkephalin. In addition, the presence of a high affinity binding site for dynorphin was demonstrated by the high potency of dynorphin (1-13) to displace [3H]EKC binding (Ki high affinity = 2.3 nM). These data provide further insights into the characterization of adrenal opioid receptors and suggest an in situ physiological role for adrenal opioid peptides.

scholarly journals The AVR4 Elicitor Protein of Cladosporium fulvum Binds to Fungal Components with High Affinity

2002 ◽  
Vol 15 (12) ◽  
pp. 1219-1227 ◽  
Author(s):  
Nienke Westerink ◽  
Ronelle Roth ◽  
Harrold A. Van den Burg ◽  
Pierre J. G. M. De Wit ◽  
Matthieu H. A. J. Joosten
Keyword(s):  
Binding Site ◽  
Specific Binding ◽  
Proteinase K ◽  
Affinity Binding ◽  
Cladosporium Fulvum ◽  
Binding Studies ◽  
Tomato Plants ◽  
High Affinity Binding ◽  
High Affinity ◽  
High Affinity Binding Site

The interaction between tomato and the fungal pathogen Cladosporium fulvum complies with the gene-for-gene system. Strains of C. fulvum that produce race-specific elicitor AVR4 induce a hypersensitive response, leading to resistance, in tomato plants that carry the Cf-4 resistance gene. The mechanism of AVR4 perception was examined by performing binding studies with 125I-AVR4 on microsomal membranes of tomato plants. We identified an AVR4 high-affinity binding site (KD = 0.05 nM) which exhibited all the characteristics expected for ligand-receptor interactions, such as saturability, reversibility, and specificity. Surprisingly, the AVR4 high-affinity binding site appeared to originate from fungi present on infected tomato plants rather than from the tomato plants themselves. Detailed analysis showed that this fungus-derived, AVR4-specific binding site is heat- and proteinase K-resistant. Affinity crosslinking demonstrated that AVR4 specifically binds to a component of approximately 75 kDa that is of fungal origin. Our data suggest that binding of AVR4 to a fungal component or components is related to the intrinsic virulence function of AVR4 for C. fulvum.

scholarly journals Characterization of the SP11/SCR High-Affinity Binding Site Involved in Self/Nonself Recognition in Brassica Self-Incompatibility

2007 ◽  
Vol 19 (1) ◽  
pp. 107-117 ◽  
Author(s):  
Hiroko Shimosato ◽  
Naohiko Yokota ◽  
Hiroshi Shiba ◽  
Megumi Iwano ◽  
Tetsuyuki Entani ◽  
...  
Keyword(s):  
Binding Site ◽  
Affinity Binding ◽  
Self Incompatibility ◽  
High Affinity Binding ◽  
High Affinity ◽  
High Affinity Binding Site ◽  
Nonself Recognition
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