Structural and Functional Characterization of the Dimerization Region of Soluble Guanylyl Cyclase

Zhou, Zongmin; Gross, Steffen; Roussos, Charis; Meurer, Sabine; Müller‐Esterl, Werner; Papapetropoulos, Andreas

doi:10.1074/jbc.m402105200

Cited by 38 publications

(44 citation statements)

References 34 publications

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“…4A), suggestive of a similar dimerization function in sGC. The hydrophobic nature of all 5 residues comprising the central cluster of the dimerization interface (Leu 8 , Leu 13 , Phe 17 , Phe 100 , and Leu 102 ) is conserved as well suggesting that sGC␣1 and sGC␤1 H-NOXA domains could similarly heterodimerize, or homodimerize as observed in solution for the sGC␤1 H-NOXA domain (Fig. 4B).…”

Section: Resultsmentioning

confidence: 87%

“…Even less is structurally known about the H-NOXA/CC region except that it was found to contain two distinct regions key for dimerization: one comprises H-NOXA ␤1 residues 204 -244, and the second contains CC residues 379 -408 with the intervening sequences postulated to contribute to a functional binding region (17). Analogous regions in sGC␣1 have also been shown to be important for sGC activity via deletion studies (9).…”

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confidence: 99%

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PAS-mediated Dimerization of Soluble Guanylyl Cyclase Revealed by Signal Transduction Histidine Kinase Domain Crystal Structure

Sayed

Baskaran

et al. 2008

Journal of Biological Chemistry

116

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Signal transduction histidine kinases (STHK) are key for sensing environmental stresses, crucial for cell survival, and attain their sensing ability using small molecule binding domains. The N-terminal domain in an STHK from Nostoc punctiforme is of unknown function yet is homologous to the central region in soluble guanylyl cyclase (sGC), the main receptor for nitric oxide (NO). This domain is termed H-NOXA (or H-NOBA) because it is often associated with the heme-nitric oxide/oxygen binding (H-NOX) domain. A structure-function approach was taken to investigate the role of H-NOXA in STHK and sGC. We report the 2.1 Å resolution crystal structure of the dimerized H-NOXA domain of STHK, which reveals a Per-Arnt-Sim (PAS) fold. The H-NOXA monomers dimerize in a parallel arrangement juxtaposing their N-terminal helices and preceding residues. Such PAS dimerization is similar to that previously observed for EcDOS, AvNifL, and RmFixL. Deletion of 7 N-terminal residues affected dimer organization. Alanine scanning mutagenesis in sGC indicates that the H-NOXA domains of sGC could adopt a similar dimer organization. Although most putative interface mutations did decrease sGC␤1 H-NOXA homodimerization, heterodimerization of full-length heterodimeric sGC was mostly unaffected, likely due to the additional dimerization contacts of sGC in the coiled-coil and catalytic domains. Exceptions are mutations sGC␣1 F285A and sGC␤1 F217A, which each caused a drastic drop in NO stimulated activity, and mutations sGC␣1 Q368A and sGC␤1 Q309A, which resulted in both a complete lack of activity and heterodimerization. Our structural and mutational results provide new insights into sGC and STHK dimerization and overall architecture.The ability to sense small molecules is key for every life form and provides information about the extracellular milieu, monitors intracellular physiological status, or establishes cell-cell communication. Sensory signaling proteins are often modular in nature with distinct domains for ligand sensing and for output signals. A number of these domains are conserved in bacteria and animals. A striking example is the heme-nitric oxide/ oxygen-binding (H-NOX) 2 (previously termed H-NOB) domain that can be a stand-alone protein in Nostoc cyanobacteria, or can be part of a multidomain protein such as in the mammalian soluble guanylyl cyclase (sGC) (1, 2) (Fig. 1A). An additional evolutionary relationship was detected between sGC and 2 other cyanobacterial signaling proteins (1, 2): the H-NOX associated H-NOXA (or H-NOBA) domain in sGC is also present at the N terminus of a cyanobacterial signal transduction histidine kinase (STHK) and 2-component hybrid sensor and regulator (2-CHSR) (Fig. 1) postulated to have a PAS-like fold (1). Both genes of these cyanobacterial proteins are adjacent to genes coding for stand-alone H-NOX domains (Fig. 1A) suggesting they might work in concert. This H-NOXA evolutionary link adds to the already complex and poorly understood regulation of sGC stimulating the need to study this ancient d...

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Section: Resultsmentioning

confidence: 87%

mentioning

confidence: 99%

PAS-mediated Dimerization of Soluble Guanylyl Cyclase Revealed by Signal Transduction Histidine Kinase Domain Crystal Structure

Sayed

Baskaran

et al. 2008

Journal of Biological Chemistry

116

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“…2A) although it exhibits the binding ability to the α 1 subunit at a reduced level (Zhou et al, 2004). In a recent study, it was reported that an N-terminal binding site of bovine α 1 subunit (amino acids position 61-128) plays major important role in heterodimerization (Wagner et al, 2005).…”

Section: Identification Of N-terminal Binding Site (Nbs) and C-terminmentioning

confidence: 99%

“…Dimerization has been attributed to the central region of the soluble GC subunits mainly based on studies with the peptide receptor/membrane GCs (Wilson and Chinkers, 1996). Recently, it has been demonstrated that the dimerization region of the β 1 -subunit consists of 205 amino acid residues over the regulatory and central regions and contains a discontinuous site of 41 amino acid residues (N-terminal binding site: NBS) and 30 amino acid residues (C-terminal binding site: CBS), respectively, facilitating binding of the β 1 subunit to the α 1 subunit (Zhou et al, 2004). On the other hand, the function of the N-terminal part of the α 1 subunit has been remained to be solved, although there are several papers to demonstrate that the N-terminal part of the α 1 subunit is critical for heme-binding (Wedel et al, 1995) and that deletion of the N-terminal 259 amino acids from the α 1 subunit had no effect on the properties of the enzyme at all, and the amino acids 259-364 of the α 1 subunit represent an important functional domain for the transduction of the NO-activation signal (Koglin and Behrends, 2003).…”

Section: Introductionmentioning

confidence: 99%

Amphipathic α-Helix Mediates the Heterodimerization of Soluble Guanylyl Cyclase

Shiga

Suzuki

2005

Zoological Science

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-Soluble guanylyl cyclase (soluble GC) is an enzyme consisting of α and β subunits and catalyzes the conversion of GTP to cGMP. The formation of heterodimer (α 1 /β 1 or α 2 /β 1 ) is essential for the catalytic activity. Each subunit has been shown to comprise three functionally different parts: a C-terminal catalytic domain, a central dimerization domain, and an N-terminal regulatory domain. The central dimerization domain in the β 1 subunit, which contains an N-terminal binding site (NBS) and a C-terminal binding site (CBS), have been postulated to be responsible for the formation of α/β heterodimers. In this study, we analyzed heterodimerization by pull down assay using the affinity between histidine tag and Ni 2+ Sepharose after co-expression of various N-and C-terminally truncated α 1 mutants (FLAG tag) and β 1 wild type (histidine tag) in the baculovirus/Sf9 system, demonstrating that the CBS-like sequence of the α 1 subunit is critical for the formation heterodimer with the β 1 subunit and the NBS-like sequence of the α 1 subunit is essential for the formation of enzymatically active heterodimer, although itself was not involved in heterodimerization. The analysis of the secondary structure of the α1 subunit predicted the existence of an amphipathic α-helix in the residues 431-464.Experiments with site-directed α 1 subunit mutant proteins demonstrated that the amphipathicity of the α-helix is important for the formation of a correct active center in the dimer.

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“…As shown in Table 3, the identity of amino acid sequences of the soluble GC α 1 and α 2 subunits between O. curvinotus and O. latipes are 98.2% and 98.5%, respectively. The dimerization region of human soluble GC-β 1 extends over 205 residues of its regulatory and central domains, and two discontinuous sites of 41 and 30 residues, respectively facilitate binding of β 1 to the α 1 subunit of human soluble GC (Zhou et al, 2004).…”

Section: Discussionmentioning

confidence: 99%

Expression of guanylyl cyclase genes in medaka hybrids (Oryzias curvinotus×Oryzias latipes)

Shiga

Suzuki

2004

Comparative Biochemistry and Physiology Part B: Biochemistry an

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The Hong Kong-originated medaka fish Oryzias curvinotus expresses nine genes (OcGC1∼OcGC8 and OcGC-R2) for membrane guanylyl cyclases (membrane GCs) and three genes (OcGCS-α 1 , OcGCS-α 2 , and OcGCS-β 1 ) for soluble GC subunits. The deduced amino acid sequences of membrane GCs expressed in O. curvinotus were quite similar to those expressed in the Japanese medaka fish O. latipes, including a novel membrane GC gene, OlGC8, which was the first isolated and characterized in O. latipes.O. curvinotus was able to produce the hybrids with O. latipes irrespective of the direction of crossing, and the resulting hybrids expressed both the maternal and paternal soluble GC subunit genes, suggesting the possibility of the formation of a chimeric heterodimer in the hybrids. In the early embryogenesis of the hybrids, however, the maternal soluble GC subunit genes were expressed earlier than the paternal soluble GC subunit genes, suggesting that the maternal soluble GC subunit genes interact more effectively with maternal effector molecules such as transcription factors than with those of paternal origin.

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Structural and Functional Characterization of the Dimerization Region of Soluble Guanylyl Cyclase

Cited by 38 publications

References 34 publications

PAS-mediated Dimerization of Soluble Guanylyl Cyclase Revealed by Signal Transduction Histidine Kinase Domain Crystal Structure

PAS-mediated Dimerization of Soluble Guanylyl Cyclase Revealed by Signal Transduction Histidine Kinase Domain Crystal Structure

Amphipathic α-Helix Mediates the Heterodimerization of Soluble Guanylyl Cyclase

Expression of guanylyl cyclase genes in medaka hybrids (Oryzias curvinotus×Oryzias latipes)

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