The Kinase Homology Domain of Receptor Guanylyl Cyclase C:  ATP Binding and Identification of an Adenine Nucleotide Sensitive Site†

Biochemistry ◽  
2006 ◽  
Vol 45 (6) ◽  
pp. 1888-1898 ◽  
Author(s):  
Mahaboobi Jaleel ◽  
Sayanti Saha ◽  
Avinash R. Shenoy ◽  
Sandhya S. Visweswariah
Keyword(s):  
Guanylyl Cyclase ◽  
Adenine Nucleotide ◽  
Atp Binding ◽  
Guanylyl Cyclase C ◽  
Sensitive Site ◽  
Kinase Homology Domain ◽  
Kinase Homology

scholarly journals Phosphorylation of the Kinase Homology Domain Is Essential for Activation of the A-Type Natriuretic Peptide Receptor

1998 ◽  
Vol 18 (4) ◽  
pp. 2164-2172 ◽  
Author(s):  
Lincoln R. Potter ◽  
Tony Hunter
Keyword(s):  
Natriuretic Peptide ◽  
Guanylyl Cyclase ◽  
Receptor Activation ◽  
Dominant Negative ◽  
Cyclase Activity ◽  
Atp Binding ◽  
Natriuretic Peptide Receptor ◽  
Peptide Receptor ◽  
Kinase Homology Domain ◽  
Kinase Homology

ABSTRACT Natriuretic peptide receptor A (NPR-A) is the biological receptor for atrial natriuretic peptide (ANP). Activation of the NPR-A guanylyl cyclase requires ANP binding to the extracellular domain and ATP binding to a putative site within its cytoplasmic region. The allosteric interaction of ATP with the intracellular kinase homology domain (KHD) is hypothesized to derepress the carboxyl-terminal guanylyl cyclase catalytic domain, resulting in the synthesis of the second messenger, cyclic GMP. Here, we show that phosphorylation of the KHD is essential for receptor activation. Using a combination of phosphopeptide mapping techniques, we have identified six residues within the ATP-binding domain (S497, T500, S502, S506, S510, and T513) which are phosphorylated when NPR-A is expressed in HEK 293 cells. Mutation of any one of these Ser or Thr residues to Ala caused reductions in the receptor phosphorylation state, the number and pattern of phosphopeptides observed in tryptic maps, and ANP-dependent guanylyl cyclase activity. The reductions were not explained by decreases in NPR-A protein levels, as indicated by immunoblot analysis and determinations of cyclase activity in the presence of detergent. Conversion of Ser-497 to Ala resulted in the most dramatic decrease in cyclase activity (∼20% of wild-type activity), but conversion to an acidic residue (Glu), which mimics the charge of the phosphoserine moiety, had no effect. Simultaneous mutation of five of the phosphorylation sites to Ala resulted in a dephosphorylated receptor which was unresponsive to hormone and had potent dominant negative inhibitory activity. We conclude that phosphorylation of the KHD is absolutely required for hormone-dependent activation of NPR-A.

Conservation of the kinaselike regulatory domain is essential for activation of the natriuretic peptide receptor guanylyl cyclases

1992 ◽  
Vol 12 (6) ◽  
pp. 2581-2590
Author(s):  
K J Koller ◽  
F J de Sauvage ◽  
D G Lowe ◽  
D V Goeddel
Keyword(s):  
Natriuretic Peptide ◽  
Guanylyl Cyclase ◽  
Growth Factor Receptor ◽  
Kinase Domain ◽  
Cyclase Activity ◽  
Guanylyl Cyclases ◽  
Heat Stable ◽  
Guanylyl Cyclase Activity ◽  
Kinase Homology Domain ◽  
Kinase Homology

The natriuretic peptide receptors, NPR-A and NPR-B, are two members of the newly described class of receptor guanylyl cyclases. The kinaselike domain of these proteins is an important regulator of the guanylyl cyclase activity. To begin to understand the molecular nature of this type of regulation, we made complete and partial deletions of the kinase domain in NPR-A and NPR-B. We also made chimeric proteins in which the kinase domains of NPR-A and NPR-B were exchanged or replaced with kinase domains from structurally similar proteins. Complete deletion of the kinase homology domain in NPR-A and NPR-B resulted in constitutive activation of the guanylyl cyclase. Various partial deletions of this region produced proteins that had no ability to activate the enzyme with or without hormone stimulation. The kinase homology domain can be exchanged between the two subtypes with no effect on regulation. However, structurally similar kinaselike domains, such as from the epidermal growth factor receptor or from the heat-stable enterotoxin receptor, another member of the receptor guanylyl cyclase family, were not able to regulate the guanylyl cyclase activity correctly. These findings suggest that the kinaselike domain of NPR-A and NPR-B requires strict sequence conservation to maintain proper regulation of their guanylyl cyclase activity.

scholarly journals A Constitutively “Phosphorylated” Guanylyl Cyclase-linked Atrial Natriuretic Peptide Receptor Mutant Is Resistant to Desensitization

1999 ◽  
Vol 10 (6) ◽  
pp. 1811-1820 ◽  
Author(s):  
Lincoln R. Potter ◽  
Tony Hunter
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Guanylyl Cyclase ◽  
Phosphorylation Sites ◽  
Natriuretic Peptide Receptor ◽  
Wild Type ◽  
Peptide Receptor ◽  
Type Receptor ◽  
Kinase Homology Domain ◽  
Kinase Homology

Dephosphorylation of the natriuretic peptide receptor-A (NPR-A) is hypothesized to mediate its desensitization in response to atrial natriuretic peptide (ANP) binding. Recently, we identified six phosphorylation sites within the kinase homology domain of NPR-A and determined that the conversion of these residues to alanine abolished the ability of the receptor to be phosphorylated or to be activated by ANP and ATP. In an attempt to generate a form of NPR-A that mimics a fully phosphorylated receptor but that is resistant to dephosphorylation, we engineered a receptor variant (NPR-A-6E) containing glutamate substitutions at all six phosphorylation sites. Consistent with the known ability of negatively charged glutamate residues to substitute functionally, in some cases, for phosphorylated residues, we found that NPR-A-6E was activated 10-fold by ANP and ATP. As determined by guanylyl cyclase assays, the hormone-stimulated activity of the wild-type receptor declined over time in membrane preparations in vitro, and this loss was blocked by the serine/threonine protein phosphatase inhibitor microcystin. In contrast, the activity of NPR-A-6E was more linear with time and was unaffected by microcystin. The nonhydrolyzable ATP analogue adenosine 5′-(β,γ-imino)-triphosphate was half as effective as ATP in stimulating the wild-type receptor but was equally as potent in stimulating NPR-A-6E, suggesting that ATP is required to keep the wild-type but not 6E variant phosphorylated. Finally, the desensitization of NPR-A-6E in whole cells was markedly blunted compared with that of the wild-type receptor, consistent with its inability to shed the negative charge from its kinase homology domain via dephosphorylation. These data provide the first direct test of the requirement for dephosphorylation in guanylyl cyclase desensitization and they indicate that it is an essential component of this process.

scholarly journals Conservation of the kinaselike regulatory domain is essential for activation of the natriuretic peptide receptor guanylyl cyclases.

1992 ◽  
Vol 12 (6) ◽  
pp. 2581-2590 ◽  
Author(s):  
K J Koller ◽  
F J de Sauvage ◽  
D G Lowe ◽  
D V Goeddel
Keyword(s):  
Natriuretic Peptide ◽  
Guanylyl Cyclase ◽  
Growth Factor Receptor ◽  
Kinase Domain ◽  
Cyclase Activity ◽  
Guanylyl Cyclases ◽  
Heat Stable ◽  
Guanylyl Cyclase Activity ◽  
Kinase Homology Domain ◽  
Kinase Homology

The natriuretic peptide receptors, NPR-A and NPR-B, are two members of the newly described class of receptor guanylyl cyclases. The kinaselike domain of these proteins is an important regulator of the guanylyl cyclase activity. To begin to understand the molecular nature of this type of regulation, we made complete and partial deletions of the kinase domain in NPR-A and NPR-B. We also made chimeric proteins in which the kinase domains of NPR-A and NPR-B were exchanged or replaced with kinase domains from structurally similar proteins. Complete deletion of the kinase homology domain in NPR-A and NPR-B resulted in constitutive activation of the guanylyl cyclase. Various partial deletions of this region produced proteins that had no ability to activate the enzyme with or without hormone stimulation. The kinase homology domain can be exchanged between the two subtypes with no effect on regulation. However, structurally similar kinaselike domains, such as from the epidermal growth factor receptor or from the heat-stable enterotoxin receptor, another member of the receptor guanylyl cyclase family, were not able to regulate the guanylyl cyclase activity correctly. These findings suggest that the kinaselike domain of NPR-A and NPR-B requires strict sequence conservation to maintain proper regulation of their guanylyl cyclase activity.

scholarly journals Opposing adenine nucleotide-dependent pathways regulate guanylyl cyclase C in rat intestine.

1994 ◽  
Vol 269 (36) ◽  
pp. 22683-22690
Author(s):  
S.J. Parkinson ◽  
S.L. Carrithers ◽  
S.A. Waldman
Keyword(s):  
Guanylyl Cyclase ◽  
Adenine Nucleotide ◽  
Guanylyl Cyclase C ◽  
Rat Intestine

scholarly journals A bone overgrowth disorder due to a gain-of-function mutation in the kinase homology domain of guanylyl cyclase B, the receptor for CNP

2013 ◽  
Vol 14 (S1) ◽  
Author(s):  
Michaela Kuhn ◽  
Thomas Premsler ◽  
Ruey-Bing Yang ◽  
Thomas D Mueller ◽  
Birgit Gaßner ◽  
...  
Keyword(s):  
Guanylyl Cyclase ◽  
Gain Of Function ◽  
Kinase Homology Domain ◽  
Kinase Homology
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