Yeast Sterol C8−C7Isomerase:  Identification and Characterization of a High-Affinity Binding Site for Enzyme Inhibitors†

Biochemistry ◽  
1996 ◽  
Vol 35 (51) ◽  
pp. 16871-16878 ◽  
Author(s):  
Fabian F. Moebius ◽  
Katrin Bermoser ◽  
Raphael J. Reiter ◽  
Markus Hanner ◽  
Hartmut Glossmann
Keyword(s):  
Binding Site ◽  
Enzyme Inhibitors ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity ◽  
High Affinity Binding Site ◽  
Identification And Characterization

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 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

Structural Characterization of a Zinc High-affinity Binding Site in Rhodopsin

2009 ◽  
Vol 85 (2) ◽  
pp. 479-484 ◽  
Author(s):  
Darwin Toledo ◽  
Arnau Cordomí ◽  
Maria Grazia Proietti ◽  
Maurizio Benfatto ◽  
Luis J. del Valle ◽  
...  
Keyword(s):  
Binding Site ◽  
Structural Characterization ◽  
Affinity Binding ◽  
High Affinity Binding ◽  
High Affinity ◽  
High Affinity Binding Site
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