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Molecular basis for small molecule inhibition of G protein-coupled receptor kinases.

Homan KT, Tesmer JJ - ACS Chem. Biol. (2014)

Bottom Line: Small molecules that inhibit the protein kinase A, G, and C (AGC) family of serine/threonine kinases can exert profound effects on cell homeostasis and thereby regulate fundamental processes such as heart rate, blood pressure, and metabolism, but there is not yet a clinically approved drug in the United States selective for a member of this family.Herein, we compare crystal structures of small molecule inhibitors in complex with GRK2 to those of highly selective compounds in complex with Rho-associated coiled-coil containing kinase 1 (ROCK1), a closely related AGC kinase.This analysis suggests that reduced hydrogen-bond formation with the hinge of the kinase domain, occupation of the hydrophobic subsite, and, consequently, higher buried surface area are key drivers of potency and selectivity among GRK inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Life Sciences Institute, Departments of Pharmacology and Biological Chemistry, University of Michigan , Ann Arbor, Michigan 48109, United States.

ABSTRACT
Small molecules that inhibit the protein kinase A, G, and C (AGC) family of serine/threonine kinases can exert profound effects on cell homeostasis and thereby regulate fundamental processes such as heart rate, blood pressure, and metabolism, but there is not yet a clinically approved drug in the United States selective for a member of this family. One subfamily of AGC kinases, the G protein-coupled receptor (GPCR) kinases (GRKs), initiates the desensitization of active GPCRs. Of these, GRK2 has been directly implicated in the progression of heart failure. Thus, there is great interest in the identification of GRK2-specific chemical probes that can be further developed into therapeutics. Herein, we compare crystal structures of small molecule inhibitors in complex with GRK2 to those of highly selective compounds in complex with Rho-associated coiled-coil containing kinase 1 (ROCK1), a closely related AGC kinase. This analysis suggests that reduced hydrogen-bond formation with the hinge of the kinase domain, occupation of the hydrophobic subsite, and, consequently, higher buried surface area are key drivers of potency and selectivity among GRK inhibitors.

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Staurosporine forms hydrogen bonds with elements conserved amongmany AGC kinase active sites but allows for multiple P-loop conformations.The view is essentially the same as that in Figure 1b. Staurosporine (gray carbons) binds in nearly superimposableconformations in PKA (yellow small lobe and green large lobe, PDBID: 1STC), PKCθ(blue, PDB ID: 1XJD), RSK1 (purple, PDB ID: 2Z7R), and PDK1 (red, PDB ID: 1OKY). The same hydrogen bonds (black dashedlines) with backbone atoms or conserved side chains are formed ineach complex (PKA numbering). However, the P-loop exhibits eitheropen (PKA and PDK1) or closed conformations (PKCθ and RSK1).The AST loop is omitted for clarity. Residue numbers correspond tothose of PKA.
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fig3: Staurosporine forms hydrogen bonds with elements conserved amongmany AGC kinase active sites but allows for multiple P-loop conformations.The view is essentially the same as that in Figure 1b. Staurosporine (gray carbons) binds in nearly superimposableconformations in PKA (yellow small lobe and green large lobe, PDBID: 1STC), PKCθ(blue, PDB ID: 1XJD), RSK1 (purple, PDB ID: 2Z7R), and PDK1 (red, PDB ID: 1OKY). The same hydrogen bonds (black dashedlines) with backbone atoms or conserved side chains are formed ineach complex (PKA numbering). However, the P-loop exhibits eitheropen (PKA and PDK1) or closed conformations (PKCθ and RSK1).The AST loop is omitted for clarity. Residue numbers correspond tothose of PKA.

Mentions: The natural product staurosporine (Figure 2a) was initially identified as a potent inhibitorof PKC (IC50 = 2 nM).39 However,even in this initial report it was noted that staurosporine exhibitscomparable inhibition of PKA and is now known as an efficacious inhibitorof many protein kinases. The crystal structure of staurosporine incomplex with PKA40 (Figure 3) demonstrated that it binds in the ATP binding site despiteexhibiting a noncompetitive mechanism of inhibition.41 The large aromatic ring system of staurosporine drivesthe two lobes of the kinase domain into a conformation slightly moreopen than the activated state, as it does when in complex with PKC,42 p90 ribosomal S6 kinase (RSK),43 and PDK1.44 The buried accessiblesurface area (ASA) for staurosporine bound to PKA is considerablyhigher than that of AMPPNP (330 vs 225 Å2, respectively),which likely underlies their difference in inhibitory potency (Table 1).


Molecular basis for small molecule inhibition of G protein-coupled receptor kinases.

Homan KT, Tesmer JJ - ACS Chem. Biol. (2014)

Staurosporine forms hydrogen bonds with elements conserved amongmany AGC kinase active sites but allows for multiple P-loop conformations.The view is essentially the same as that in Figure 1b. Staurosporine (gray carbons) binds in nearly superimposableconformations in PKA (yellow small lobe and green large lobe, PDBID: 1STC), PKCθ(blue, PDB ID: 1XJD), RSK1 (purple, PDB ID: 2Z7R), and PDK1 (red, PDB ID: 1OKY). The same hydrogen bonds (black dashedlines) with backbone atoms or conserved side chains are formed ineach complex (PKA numbering). However, the P-loop exhibits eitheropen (PKA and PDK1) or closed conformations (PKCθ and RSK1).The AST loop is omitted for clarity. Residue numbers correspond tothose of PKA.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4301174&req=5

fig3: Staurosporine forms hydrogen bonds with elements conserved amongmany AGC kinase active sites but allows for multiple P-loop conformations.The view is essentially the same as that in Figure 1b. Staurosporine (gray carbons) binds in nearly superimposableconformations in PKA (yellow small lobe and green large lobe, PDBID: 1STC), PKCθ(blue, PDB ID: 1XJD), RSK1 (purple, PDB ID: 2Z7R), and PDK1 (red, PDB ID: 1OKY). The same hydrogen bonds (black dashedlines) with backbone atoms or conserved side chains are formed ineach complex (PKA numbering). However, the P-loop exhibits eitheropen (PKA and PDK1) or closed conformations (PKCθ and RSK1).The AST loop is omitted for clarity. Residue numbers correspond tothose of PKA.
Mentions: The natural product staurosporine (Figure 2a) was initially identified as a potent inhibitorof PKC (IC50 = 2 nM).39 However,even in this initial report it was noted that staurosporine exhibitscomparable inhibition of PKA and is now known as an efficacious inhibitorof many protein kinases. The crystal structure of staurosporine incomplex with PKA40 (Figure 3) demonstrated that it binds in the ATP binding site despiteexhibiting a noncompetitive mechanism of inhibition.41 The large aromatic ring system of staurosporine drivesthe two lobes of the kinase domain into a conformation slightly moreopen than the activated state, as it does when in complex with PKC,42 p90 ribosomal S6 kinase (RSK),43 and PDK1.44 The buried accessiblesurface area (ASA) for staurosporine bound to PKA is considerablyhigher than that of AMPPNP (330 vs 225 Å2, respectively),which likely underlies their difference in inhibitory potency (Table 1).

Bottom Line: Small molecules that inhibit the protein kinase A, G, and C (AGC) family of serine/threonine kinases can exert profound effects on cell homeostasis and thereby regulate fundamental processes such as heart rate, blood pressure, and metabolism, but there is not yet a clinically approved drug in the United States selective for a member of this family.Herein, we compare crystal structures of small molecule inhibitors in complex with GRK2 to those of highly selective compounds in complex with Rho-associated coiled-coil containing kinase 1 (ROCK1), a closely related AGC kinase.This analysis suggests that reduced hydrogen-bond formation with the hinge of the kinase domain, occupation of the hydrophobic subsite, and, consequently, higher buried surface area are key drivers of potency and selectivity among GRK inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Life Sciences Institute, Departments of Pharmacology and Biological Chemistry, University of Michigan , Ann Arbor, Michigan 48109, United States.

ABSTRACT
Small molecules that inhibit the protein kinase A, G, and C (AGC) family of serine/threonine kinases can exert profound effects on cell homeostasis and thereby regulate fundamental processes such as heart rate, blood pressure, and metabolism, but there is not yet a clinically approved drug in the United States selective for a member of this family. One subfamily of AGC kinases, the G protein-coupled receptor (GPCR) kinases (GRKs), initiates the desensitization of active GPCRs. Of these, GRK2 has been directly implicated in the progression of heart failure. Thus, there is great interest in the identification of GRK2-specific chemical probes that can be further developed into therapeutics. Herein, we compare crystal structures of small molecule inhibitors in complex with GRK2 to those of highly selective compounds in complex with Rho-associated coiled-coil containing kinase 1 (ROCK1), a closely related AGC kinase. This analysis suggests that reduced hydrogen-bond formation with the hinge of the kinase domain, occupation of the hydrophobic subsite, and, consequently, higher buried surface area are key drivers of potency and selectivity among GRK inhibitors.

Show MeSH
Related in: MedlinePlus