Characterization of the hormone binding site of natriuretic peptide receptor‐C

Abstract

Key words: Atrial natriuretic peptide; Natriuretic peptide receptor; Mutagenesis; Receptor/hormone interaction dues Cys 428 and Cys 431 (for the splice variant NPR-C6) are involved in NPR-C homodimer formation while the remaining four cysteine residues form two intramolecular loops (Cys 63-Cys 9~ and Cys168-Cys216). In a previous paper we reported the sequence analysis and functional expression of rat NPR-C, which shows a higher affinity to all natriuretic peptides compared to the human receptor [9]. Orthologous mutagenesis at position 188 for rat and human NPR-C results in a complete reversal of the pharmacology of both receptors [9]. In this paper we report the detection of a second residue at position 205 with a more specific influence on binding to a certain number of naturally occurring and synthetic ligands. Double mutations as well as alanine mutations at positions 188 and 205 further delineate differences in hormone binding between rat and human NPR-C.

Structural Determinants of Natriuretic Peptide Receptor Specificity and Degeneracy

“…As mentioned, mutation of Ile188 to Ala results in a higher affinity for NP hormones, and substituting residues 188 and 205 from human NPR-C with rat, results in a narrowing of ANP analog specificity, as seen in the rat receptor. 28 Mapping the conservation of various NPRs onto the NPR-C structure ( Figure 5(b)) also supports the notion that pocket I is important for specificity: pocket I shows the most volume and shape differences between NPR-C and NPR-A, and appears less conserved. The greater sequence variation in the lower NP-binding region therefore may be correlated with ligand specificity determinants.…”

“…Importantly, increasing the volume of NPR-C pocket I by mutation of Ile188 to Ala results in higher affinity for NP hormones. 28 Although ANP, BNP and CNP all place a phenylalanine in this pocket, the structural differences between ANP/BNP and CNP indicates that the NPR-A pocket I has a more optimal fit with the Phe8 of ANP/BNP than it would have with Phe7 of CNP. As shown in Figure 2(d), CNP adopts a more extended main chain conformation than ANP/BNP, which in turn results in the CNP Phe7 extending further away from the center of the ring and deeper into pocket I.…”

“…Mutagenesis studies have shown that Ile188 and Asn205 of NPR-C are important for modulating the ligand specificity of this receptor. 28 Ile188 is a critical component of pocket I; Asn205 is directly below the pocket, which may have interactions with the N and/or C-terminal residues of NPs where the sequences show significant differences. As mentioned, mutation of Ile188 to Ala results in a higher affinity for NP hormones, and substituting residues 188 and 205 from human NPR-C with rat, results in a narrowing of ANP analog specificity, as seen in the rat receptor.…”

Expression and Genomic Organization of A Medaka Fish Novel Membrane Form of Guanylyl Cyclase/Orphan Receptor

“…To the best of our knowledge, no paper has yet demonstrated the activation of any type of natriuretic peptide receptor by C-ANF. Along these lines, OlGC7 is known to be a unique natriuretic peptide receptor; the activation of OlGC7 by C-ANF and various medaka fish tissue extracts suggests that the extracellular domain of OlGC7 is structurally similar to that of NPRC, although the residues Ile 188 and Asn 205 in the extracellular domain of human NPRC, which are important for modulating hormone specificity (Engel and Lowe, 1995), are not conserved in the extracellular domain of OlGC7. It is known that the Japanese eel Anguilla japonica contains ANP and CNP and a unique natriuretic peptide, VNP, instead of BNP; it also contains two known natriuretic peptide receptors, NPR-A (GC-A) and NPR-B (GC-B), and natriuretic peptide clearance receptors (NPR-C and NPR-D) (Takei and Hirose, 2002).…”

Mutations in B‐type natriuretic peptide mediating receptor‐A selectivity

“…These data highlight fundamental differences in the mode of ligand recognition by NPR-C and NPR-A. This conclusion is also true for cross-species comparisons of NPR-A and NPR-C, with divergent modes of recognition for both ANP and BNP variants [14][15][16]. Together these observations suggest that each receptor/ligand pair will have some unique structural determinants of specificity and affinity, perhaps reflected in a variable contour and different side-chain contacts at the receptor/ ligand interface.…”