Proper glycosylation and phosphorylation of the type A natriuretic peptide receptor are required for hormone-stimulated guanylyl cyclase activity.
The natriuretic peptide receptor type A (NPR-A) is a receptor-guanylyl cyclase whose cytoplasmic enzymatic activity is stimulated by atrial natriuretic peptide binding to the extracellular domain. NPR-A expressed in COS cells is heterogeneously glycosylated, and the more highly glycosylated protein is also phosphorylated. Upon hormone binding, dephosphorylation occurs from both serine and threonine residues, probably within the kinase homology domain of NPR-A, and may be involved with receptor desensitization. Using site-specific mutations in the kinase homology domain of NPR-A, we have identified several residues that are important for regulating the guanylyl cyclase activity of NPR-A. Some of these amino acids are probably essential for maintaining the proper tertiary structure of the intracellular domain, and others may form loops that allow for binding of ATP, which is required for proper enzymatic activity. The site-specific mutants which have greatly reduced enzymatic activity are not phosphorylated and are incompletely glycosylated. These results suggest a correlation between phosphorylation and complete glycosylation of NPR-A and that both are required for hormone-induced enzymatic activity.
Koller
K J
KJ
Department of Molecular Biology, Genentech Inc., South San Francisco, California 94080.
Lipari
M T
MT
Goeddel
D V
DV
D000595:Amino Acid Sequence; D000818:Animals; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D001483:Base Sequence; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004277:DNA, Single-Stranded; D004789:Enzyme Activation; D006031:Glycosylation; D006162:Guanylate Cyclase / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D017461:Receptors, Atrial Natriuretic Factor / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D004277:DNA, Single-Stranded; D011994:Recombinant Proteins; D009320:Atrial Natriuretic Factor; D011494:Protein Kinases; D006162:Guanylate Cyclase; D017461:Receptors, Atrial Natriuretic Factor