Abstract:
:Activation of different forms of adenylate cyclases (AC) by forskolin and displacement of [14,15-3H]dihydroforskolin binding from membranes by forskolin in the absence or presence of specific stimulatory hormone and beta, gamma-imidoguanosine 5'-triphosphate (Gpp(NH)p) have been studied. These conditions have been used to generate forskolin dose-response curves of AC activation. A plot of enzyme activation versus apparent forskolin-binding showed a linear and a nonlinear relationship, respectively, in the absence or presence of the other two stimulators. The latter relationship can be fitted by two linear regression lines with a defined intercept, the slopes of which represent two distinct binding-activation (B-A) effects. The B-A effects of forskolin for rat adipocyte and liver membranes in the absence of stimulatory hormone and Gpp(NH)p were 10 and 8 (pmol X min-1) X (pmol)-1, respectively. The B-A effects for the same membranes in the presence of the other two stimulators were 69 (high) and 13 (low) (pmol X min-1) X (pmol)-1 for adipocyte membrane, and 83 (high) and 9 (low) (pmol X min-1) X (pmol)-1 for liver membrane. The ratio of potentiation of forskolin-activated enzyme activity to the unmodified forskolin-stimulated activity (P-A ratio) was determined without the binding data. At 3 microM forskolin, with and without 230 epinephrine and 10 microM Gpp(NH)p, the P-A ratio was 3.7, decreasing to 1.1 with the addition 100 microM forskolin. The line representing a high B-A effect and a resulting high P-A ratio appears to describe the interactions between forskolin and the AC stimulated by epinephrine and Gpp(NH)p. The line of low B-A effect may represent the interaction between forskolin and the basal AC. Two peaks of AC activity were eluted from forskolin-Sepharose column. They have apparent differences in sensitivity to Gpp(NH)p and affinity for forskolin. Based on the results available thus far, with consideration for known limitations of the methodology, a working model has been proposed for forskolin activation of AC.
journal_name
Arch Biochem Biophysjournal_title
Archives of biochemistry and biophysicsauthors
Shi QH,Ruiz JA,Ho RJdoi
10.1016/0003-9861(86)90062-7subject
Has Abstractpub_date
1986-11-15 00:00:00pages
156-65issue
1eissn
0003-9861issn
1096-0384pii
0003-9861(86)90062-7journal_volume
251pub_type
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