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dc.contributor.authorPinto, Cibele
dc.contributor.authorHubner, Melanie
dc.contributor.authorGille, Andreas
dc.contributor.authorRichter, Mark
dc.contributor.authorMou, Tung-Chung
dc.contributor.authorSprang, Stephen R.
dc.contributor.authorSeifert, Roland
dc.date.accessioned2017-01-27T20:04:42Z
dc.date.available2017-01-27T20:04:42Z
dc.date.issued2009-04-02
dc.identifier.citationPinto, Cibele, Melanie Hübner, Andreas Gille, Mark Richter, Tung-Chung Mou, Stephen R. Sprang, and Roland Seifert. "Differential Interactions of the Catalytic Subunits of Adenylyl Cyclase with Forskolin Analogs." Biochemical Pharmacology 78.1 (2009): 62-69.en_US
dc.identifier.urihttp://hdl.handle.net/1808/22681
dc.description.abstractThe diterpene forskolin (FS) binds to, and activates, mammalian membranous adenylyl cyclase (AC) isoforms I–VIII. Diterpenes without C1-OH group do not activate ACs. The C1-OH group forms a hydrogen bond with the backbone oxygen of Val506 of the C1 catalytic subunit of AC (isoform V numbering). To better understand the mechanism of AC activation we examined the interactions of FS and eight FS analogs with purified catalytic AC subunits C1 (AC V) and C2 (AC II) by fluorescence spectroscopy, using 2′,3′-O-(N-methylanthraniloyl)-guanosine 5′-triphosphate (MANT-GTP) as fluorescent reporter probe, and by enzymatic activity. FS analogs induced C1/C2 assembly as assessed by fluorescence resonance energy transfer from Trp1020 of C2 to MANT-GTP and by increased direct MANT-GTP fluorescence in the order of efficacy FS ~ 7-deacetyl-FS ~ 6-acetyl-7-deacetyl-FS ~ 9-deoxy-FS > 7-deacetyl-7-(N-methylpiperazino-γ-butyryloxy)-FS > 1-deoxy-FS ~ 1,9-dideoxy-FS ~ 7-deacetyl-1-deoxy-FS ~ 7-deacetyl-1,9-dideoxy-FS. In contrast, FS analogs activated catalysis in the order of efficacy FS > 7-deacety-FS ~ 6-acetyl-7-deacetyl-FS ~ 9-deoxy-FS > 7-deacetyl-7-(N-methylpiperazino-γ-butyryloxy)-FS ≫ 1-deoxy-FS, 1,9-dideoxy-FS, 7-deacetyl-1-deoxy-FS and 7-deacetyl-1,9-dideoxy-FS (all ineffective). 1-Deoxy-FS analogs inhibited FS-stimulated catalysis by an apparently non-competitive mechanism. Our data suggest a two-step mechanism of AC activation by diterpenes. In the first step, diterpenes, regardless of their substitution pattern, promote C1/C2 assembly. In the second and yet poorly understood step, diterpenes that form a hydrogen bond between C1-OH and Val506 promote a conformational switch that results in activation of catalysis. The apparent non-competitive interaction of FS with 1-deoxy-FS analogs is explained by impaired ligand exchange due to strong hydrophobic interactions with C1/C2.en_US
dc.publisherElsevieren_US
dc.rightsThis is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License 3.0 (CC BY-NC-ND 3.0 US), which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.en_US
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/3.0/
dc.titleDifferential Interactions of the Catalytic Subunits of Adenylyl Cyclase with Forskolin Analogsen_US
dc.typeArticleen_US
kusw.kuauthorRichter, Mark
kusw.kudepartmentMolecular Biosciencesen_US
dc.identifier.doi10.1016/j.bcp.2009.03.023en_US
kusw.oaversionScholarly/refereed, author accepted manuscripten_US
kusw.oapolicyThis item meets KU Open Access policy criteria.en_US
dc.rights.accessrightsopenAccess


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This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License 3.0 (CC BY-NC-ND 3.0 US), which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.
Except where otherwise noted, this item's license is described as: This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License 3.0 (CC BY-NC-ND 3.0 US), which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.