1U0H

STRUCTURAL BASIS FOR THE INHIBITION OF MAMMALIAN ADENYLYL CYCLASE BY MANT-GTP

Summary for 1U0H

• Entry DOI: 10.2210/pdb1u0h/pdb
• Related: 1TL7
• Descriptor: Adenylate cyclase, type V, Adenylate cyclase, type II, Guanine nucleotide-binding protein G(s), alpha subunit, ... (9 entities in total)
• Functional Keywords: adenylyl cyclase, gsa, mant-gtp, lyase
• Biological source: Canis lupus familiaris (dog); More
• Cellular location: Membrane; Multi-pass membrane protein: P30803 P26769; Cell membrane; Lipid-anchor (By similarity): P04896
• Total number of polymer chains: 3
• Total formula weight: 96727.52

Authors

Mou, T.C.,Gille, A.,Seifert, R.J.,Sprang, S.R. (deposition date: 2004-07-13, release date: 2004-12-14, Last modification date: 2023-08-23)

Primary citation

Mou, T.C.,Gille, A.,Fancy, D.A.,Seifert, R.,Sprang, S.R.
Structural basis for the inhibition of mammalian membrane adenylyl cyclase by 2 '(3')-O-(N-Methylanthraniloyl)-guanosine 5 '-triphosphate.
J.Biol.Chem., 280:7253-7261, 2005

PubMed Abstract: Membrane-bound mammalian adenylyl cyclase (mAC) catalyzes the synthesis of intracellular cyclic AMP from ATP and is activated by stimulatory G protein alpha subunits (Galpha(s)) and by forskolin (FSK). mACs are inhibited with high potency by 2 '(3')-O-(N-methylanthraniloyl) (MANT)-substituted nucleotides. In this study, the crystal structures of the complex between Galpha(s).GTPgammaS and the catalytic C1 and C2 domains from type V and type II mAC (VC1.IIC2), bound to FSK and either MANT-GTP.Mg(2+) or MANT-GTP.Mn(2+) have been determined. MANT-GTP coordinates two metal ions and occupies the same position in the catalytic site as P-site inhibitors and substrate analogs. However, the orientation of the guanine ring is reversed relative to that of the adenine ring. The MANT fluorophore resides in a hydrophobic pocket at the interface between the VC1 and IIC2 domains and prevents mAC from undergoing the "open" to "closed" domain rearrangement. The K(i) of MANT-GTP for inhibition of VC1.IIC2 is lower in the presence of mAC activators and lower in the presence of Mn(2+) compared with Mg(2+), indicating that the inhibitor binds more tightly to the catalytically most active form of the enzyme. Fluorescence resonance energy transfer-stimulated emission from the MANT fluorophore upon excitation of Trp-1020 in the MANT-binding pocket of IIC2 is also stronger in the presence of FSK. Mutational analysis of two non-conserved amino acids in the MANT-binding pocket suggests that residues outside of the binding site influence isoform selectivity toward MANT-GTP.

PubMed: 15591060
DOI: 10.1074/jbc.M409076200

Experimental method

X-RAY DIFFRACTION (2.9 Å)