Abstract
The inhibitory interaction of phosphodiesterase-6 (PDE6) with its gamma-subunit (Pgamma) is pivotal in vertebrate phototransduction. Here, crystal structures of a chimaeric PDE5/PDE6 catalytic domain (PDE5/6cd) complexed with sildenafil or 3-isobutyl-1-methylxanthine and the Pgamma-inhibitory peptide Pgamma(70-87) have been determined at 2.9 and 3.0 A, respectively. These structures show the determinants and the mechanism of the PDE6 inhibition by Pgamma and suggest the conformational change of Pgamma on transducin activation. Two variable H- and M-loops of PDE5/6cd form a distinct interface that contributes to the Pgamma-binding site. This allows the Pgamma C-terminus to fit into the opening of the catalytic pocket, blocking cGMP access to the active site. Our analysis suggests that disruption of the H-M loop interface and Pgamma-binding site is a molecular cause of retinal degeneration in atrd3 mice. Comparison of the two PDE5/6cd structures shows an overlap between the sildenafil and Pgamma(70-87)-binding sites, thereby providing critical insights into the side effects of PDE5 inhibitors on vision.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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1-Methyl-3-isobutylxanthine / chemistry
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1-Methyl-3-isobutylxanthine / metabolism
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1-Methyl-3-isobutylxanthine / pharmacology
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Amino Acid Sequence
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Animals
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Catalytic Domain
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Cattle
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Cyclic Nucleotide Phosphodiesterases, Type 5 / chemistry
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Cyclic Nucleotide Phosphodiesterases, Type 5 / metabolism
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Cyclic Nucleotide Phosphodiesterases, Type 6 / antagonists & inhibitors*
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Cyclic Nucleotide Phosphodiesterases, Type 6 / chemistry*
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Cyclic Nucleotide Phosphodiesterases, Type 6 / metabolism
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Cyclic Nucleotide Phosphodiesterases, Type 6 / physiology
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Enzyme Inhibitors / chemistry
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Enzyme Inhibitors / metabolism
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Enzyme Inhibitors / pharmacology
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Humans
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Models, Molecular
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Molecular Sequence Data
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Peptide Fragments / chemistry
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Peptide Fragments / metabolism
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Peptide Fragments / pharmacology
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Piperazines / chemistry
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Piperazines / metabolism
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Piperazines / pharmacology
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Protein Binding
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Protein Subunits / chemistry
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Protein Subunits / physiology
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Purines / chemistry
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Purines / metabolism
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Purines / pharmacology
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / metabolism
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Sequence Homology, Amino Acid
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Sildenafil Citrate
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Structure-Activity Relationship
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Sulfones / chemistry
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Sulfones / metabolism
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Sulfones / pharmacology
Substances
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Enzyme Inhibitors
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Peptide Fragments
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Piperazines
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Protein Subunits
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Purines
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Recombinant Fusion Proteins
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Sulfones
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Sildenafil Citrate
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Cyclic Nucleotide Phosphodiesterases, Type 5
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Cyclic Nucleotide Phosphodiesterases, Type 6
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PDE6G protein, human
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1-Methyl-3-isobutylxanthine