Role of the nucleotidyl cyclase helical domain in catalytically active dimer formation

Vercellino, Irene; Rezabkova, Lenka; Oliéric, V.; Polyhach, Yevhen; Weinert, T.; Kammerer, Richard A.; Jeschke, Gunnar; Korkhov, Volodymyr M.

doi:10.1073/pnas.1712621114

Cited by 40 publications

(57 citation statements)

References 46 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Except for small single-domain globular proteins, one may need to test whether a crystal structure is representative of the solution structure. PDS-based distance distributions can provide this information [70], which is particularly useful when different techniques result in different predictions of solution structure [71] or when a shortened construct had to be used for crystallization [46,72].…”

Section: Integrative Modellingmentioning

confidence: 99%

The contribution of modern EPR to structural biology

Jeschke

2018

Emerging Topics in Life Sciences

Self Cite

112

133

View full text Add to dashboard Cite

Electron paramagnetic resonance (EPR) spectroscopy combined with site-directed spin labelling is applicable to biomolecules and their complexes irrespective of system size and in a broad range of environments. Neither short-range nor long-range order is required to obtain structural restraints on accessibility of sites to water or oxygen, on secondary structure, and on distances between sites. Many of the experiments characterize a static ensemble obtained by shock-freezing. Compared with characterizing the dynamic ensemble at ambient temperature, analysis is simplified and information loss due to overlapping timescales of measurement and system dynamics is avoided. The necessity for labelling leads to sparse restraint sets that require integration with data from other methodologies for building models. The double electron-electron resonance experiment provides distance distributions in the nanometre range that carry information not only on the mean conformation but also on the width of the native ensemble. The distribution widths are often inconsistent with Anfinsen's concept that a sequence encodes a single native conformation defined at atomic resolution under physiological conditions.

show abstract

Section: Integrative Modellingmentioning

confidence: 99%

The contribution of modern EPR to structural biology

Jeschke

2018

Emerging Topics in Life Sciences

Self Cite

112

133

View full text Add to dashboard Cite

show abstract

“…30,31 Structures of membrane-bound adenylate cyclases have been solved that contain catalytic domains as well as portions of or the entire CC domains, helping to orient the C-terminal domains of sGC. 32,33 However, the precise quaternary structural arrangement of domains in the N-terminal portion of sGC is not known. Consequently, the mechanism by which NO and small-molecule stimulators couple binding through the protein for activation are poorly understood.…”

Section: Introductionmentioning

confidence: 99%

Allosteric activation of the nitric oxide receptor soluble guanylate cyclase mapped by cryo-electron microscopy

Horst

Yokom

Rosenberg

et al. 2019

Preprint

View full text Add to dashboard Cite

24Soluble guanylate cyclase (sGC) is the primary receptor for nitric oxide (NO) in 25 mammalian nitric oxide signaling. We determined structures of full-length Manduca sexta 26 sGC in both inactive and active states using cryo-electron microscopy. NO and the sGC-27 specific stimulator YC-1 induce a 71° rotation of the heme-binding β H-NOX and PAS 28 domains. Repositioning of the β H-NOX domain leads to a straightening of the coiled-coil 29 domains, which, in turn, use the motion to move the catalytic domains into an active 30 conformation. YC-1 binds directly between the β H-NOX domain and the two CC domains. 31 The structural elongation of the particle observed in cryo-EM was corroborated in solution 32 using small angle X-ray scattering (SAXS). These structures delineate the endpoints of 33 the allosteric transition responsible for the major cyclic GMP-dependent physiological 34 effects of NO. 36Nitric oxide (NO) is a critical primary signaling molecule in eukaryotic organisms. 1,2 38Cells detect this diatomic gas through the NO receptor soluble guanylate cyclase (sGC), 39 which is activated by NO to catalyze the cyclization of 5′-guanosine triphosphate (GTP) 40 to 3′,5′-cyclic guanosine monophosphate (cGMP). [3][4][5][6][7][8] The NO signaling pathway, with 41 sGC as a central component, plays crucial roles in the cardiovascular and neurological 42 systems in mammals. 9-11 Furthermore, aberrations in these signaling pathways can lead 43 to pathologies that include various forms of hypertension, cardiovascular disease, and 44 neurodegeneration. 12-16 45 sGC is a heterodimer composed of two subunits, denoted α and β, that are each 46 composed of four domains: an N-terminal heme nitric oxide/oxygen (H-NOX) domain, a 47 Per/Arnt/Sim (PAS)-like domain, a coiled-coil (CC) domain, and a catalytic (CAT) 48 domain. 6,7 Although each subunit contains an H-NOX domain, only the β H-NOX domain 49 binds a heme cofactor, with direct ligation occurring through a conserved histidine 50 residue. 17 The CC domains, together with the PAS domains, are thought to form a 51 structured assembly upon dimerization of sGC. 18 The CAT domains form a wreath-like 52 structure with the active site at the dimer interface. 6,19 53Biochemical aspects of sGC activation by NO have been studied in great detail. 54 Without a ligand bound to the β H-NOX domain, sGC has a low basal activity. A 55 stoichiometric equivalent of NO relative to the sGC heterodimer results in the cleavage of 56 the proximal histidine-iron bond and the formation of a distal five-coordinate ferrous 57 nitrosyl enzyme with 15% of maximal activity. 5,20-22 This low-activity state of sGC will be 58 referred to here as the 1-NO state; importantly, the KD of the ferrous nitrosyl heme is 1.2 59 4× 10 -12 M thus in this state NO remains bound to the heme. 23 The activity of the 1-NO 60 state can be increased to a maximally-active state either by the addition of excess NO 61 (xsNO), or by addition of small-molecule stimulators (e.g. YC-1). 24,25 The molecular 62 mechanis...

show abstract

“…This interpretation was subsequently confirmed through analysis of chemical cross‐links in sGC that were only consistent with a parallel helical arrangement, and through single‐particle electron microscopy . The arrangement is also found in adenyl cyclase, which is evolutionarily related to guanylyl cyclase and retains a short coiled coil attached to the cyclase domain through a helix‐turn‐helix motif, as anticipated to occur in sGC . The sGC coiled coil is a central domain connecting the NO‐sensing half of the sGC heterodimer to the catalytic domains.…”

Section: Introductionmentioning

confidence: 99%

“…25 The arrangement is also found in adenyl cyclase, which is evolutionarily related to guanylyl cyclase and retains a short coiled coil attached to the cyclase domain through a helix-turn-helix motif, as anticipated to occur in sGC. 26,27 The sGC coiled coil is a central domain connecting the NO-sensing half of the sGC heterodimer to the catalytic domains. It also serves as a noncovalent dimerization element for heterodimer assembly and may function as a signaling helix.…”

Section: Introductionmentioning

confidence: 99%

Instability in a coiled‐coil signaling helix is conserved for signal transduction in soluble guanylyl cyclase

et al. 2019

View full text Add to dashboard Cite

How nitric oxide (NO) activates its primary receptor, α1/β1 soluble guanylyl cyclase (sGC or GC‐1), remains unknown. Likewise, how stimulatory compounds enhance sGC activity is poorly understood, hampering development of new treatments for cardiovascular disease. NO binding to ferrous heme near the N‐terminus in sGC activates cyclase activity near the C‐terminus, yielding cGMP production and physiological response. CO binding can also stimulate sGC, but only weakly in the absence of stimulatory small‐molecule compounds, which together lead to full activation. How ligand binding enhances catalysis, however, has yet to be discovered. Here, using a truncated version of sGC from Manduca sexta, we demonstrate that the central coiled‐coil domain, the most highly conserved region of the ~150,000 Da protein, not only provides stability to the heterodimer but is also conformationally active in signal transduction. Sequence conservation in the coiled coil includes the expected heptad‐repeating pattern for coiled‐coil motifs, but also invariant positions that disfavor coiled‐coil stability. Full‐length coiled coil dampens CO affinity for heme, while shortening of the coiled coil leads to enhanced CO binding. Introducing double mutation αE447L/βE377L, predicted to replace two destabilizing glutamates with leucines, lowers CO binding affinity while increasing overall protein stability. Likewise, introduction of a disulfide bond into the coiled coil results in reduced CO affinity. Taken together, we demonstrate that the heme domain is greatly influenced by coiled‐coil conformation, suggesting communication between heme and catalytic domains is through the coiled coil. Highly conserved structural imperfections in the coiled coil provide needed flexibility for signal transduction.

show abstract

Role of the nucleotidyl cyclase helical domain in catalytically active dimer formation

Cited by 40 publications

References 46 publications

The contribution of modern EPR to structural biology

The contribution of modern EPR to structural biology

Allosteric activation of the nitric oxide receptor soluble guanylate cyclase mapped by cryo-electron microscopy

Instability in a coiled‐coil signaling helix is conserved for signal transduction in soluble guanylyl cyclase

Contact Info

Product

Resources

About