Structural Perspectives on the Mechanism of Soluble Guanylate Cyclase Activation

Wittenborn, E.C.; Marletta, Michael A.

doi:10.3390/ijms22115439

Cited by 25 publications

(20 citation statements)

References 74 publications

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“…Conventional sGC activation proceeds through NO binding to a ferrous heme located within the sGCβ subunit of a mature sGC α–β heterodimer ( 13 , 14 ). This causes protein structural changes that activate cGMP production ( 13 – 16 ). The essential role that heme plays in sensing NO underscores the importance of the heme delivery and insertion steps that enable sGC to mature in cells ( 17 ).…”

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confidence: 99%

NO rapidly mobilizes cellular heme to trigger assembly of its own receptor

Dai

Faul

Ghosh

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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Nitric oxide (NO) signaling in biology relies on its activating cyclic guanosine monophosphate (cGMP) production by the NO receptor soluble guanylyl cyclase (sGC). sGC must obtain heme and form a heterodimer to become functional, but paradoxically often exists as an immature heme-free form in cells and tissues. Based on our previous finding that NO can drive sGC maturation, we investigated its basis by utilizing a fluorescent sGC construct whose heme level can be monitored in living cells. We found that NO generated at physiologic levels quickly triggered cells to mobilize heme to immature sGC. This occurred when NO was generated within cells or by neighboring cells, began within seconds of NO exposure, and led cells to construct sGC heterodimers and thus increase their active sGC level by several-fold. The NO-triggered heme deployment involved cellular glyceraldehyde-3-phosphate dehydrogenase (GAPDH)–heme complexes and required the chaperone hsp90, and the newly formed sGC heterodimers remained functional long after NO generation had ceased. We conclude that NO at physiologic levels triggers assembly of its own receptor by causing a rapid deployment of cellular heme. Redirecting cellular heme in response to NO is a way for cells and tissues to modulate their cGMP signaling and to more generally tune their hemeprotein activities wherever NO biosynthesis takes place.

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confidence: 99%

NO rapidly mobilizes cellular heme to trigger assembly of its own receptor

Dai

Faul

Ghosh

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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“…The 3-D structure of the sGC β-subunit H-NOX domain is remarkably well-conserved among species, while the amino acids of the H-NOX cavity which mediate the interaction of H-NOX with heme are remarkably well-conserved between bacterial and mammalian species (e.g. Nostoc punctiforme, Bos taurus ), being identical by ∼63% (17 of 27, ( Alexandropoulos et al., 2016 ; Makrynitsa et al., 2019 ; Wittenborn and Marletta, 2021 ). This is why rational drug design based on microbial H-NOX structural data and on the very structure of heme has contributed to, and still informs, the development of sGC activator molecules all the way to advanced clinical trials (reviewed in Alexandropoulos et al., 2016 ; Makrynitsa et al., 2019 ; Liu et al., 2021 ; Wittenborn and Marletta, 2021 ).…”

Section: Resultsmentioning

confidence: 99%

“…Nostoc punctiforme, Bos taurus ), being identical by ∼63% (17 of 27, ( Alexandropoulos et al., 2016 ; Makrynitsa et al., 2019 ; Wittenborn and Marletta, 2021 ). This is why rational drug design based on microbial H-NOX structural data and on the very structure of heme has contributed to, and still informs, the development of sGC activator molecules all the way to advanced clinical trials (reviewed in Alexandropoulos et al., 2016 ; Makrynitsa et al., 2019 ; Liu et al., 2021 ; Wittenborn and Marletta, 2021 ). Despite all this, it is understood that ideally, any conclusions derived from a structural approach using a recombinant microbial domain (in this case Nostoc sp.…”

Section: Resultsmentioning

confidence: 99%

Replacement of heme by soluble guanylate cyclase (sGC) activators abolishes heme-nitric oxide/oxygen (H-NOX) domain structural plasticity

Argyriou

Makrynitsa

Δάλκας

et al. 2021

Current Research in Structural Biology

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The gasotransmitter nitric oxide (NO) is a critical endogenous regulator of homeostasis, in major part via the generation of cGMP (cyclic guanosine monophosphate) from GTP (guanosine triphosphate) by NO's main physiological receptor, the soluble guanylate cyclase (sGC). sGC is a heterodimer, composed of an α1 and a β1 subunit, of which the latter contains the heme-nitric oxide/oxygen (H-NOX) domain, responsible for NO recognition, binding and signal initiation. The NO/sGC/cGMP axis is dysfunctional in a variety of diseases, including hypertension and heart failure, especially since oxidative stress results in heme oxidation, sGC unresponsiveness to NO and subsequent degradation. As a central player in this axis, sGC is the focus of intense research efforts aiming to develop therapeutic molecules that enhance its activity. A class of drugs named sGC “activators” aim to replace the oxidized heme of the H-NOX domain, thus stabilizing the enzyme and restoring its activity. Although numerous studies outline the pharmacology and binding behavior of these compounds, the static 3D models available so far do not allow a satisfactory understanding of the structural basis of sGC's activation mechanism by these drugs. Herein, application NMR describes different conformational states during the replacement of the heme by a sGC activators. We show that the two sGC activators (BAY 58-2667 and BAY 60-2770) significantly decrease the conformational plasticity of the recombinant H-NOX protein domain of Nostoc sp. cyanobacterium, rendering it a lot more rigid compared to the heme-occupied H-NOX. NMR methodology also reveals, for the first time, a surprising bi-directional competition between reduced heme and these compounds, pointing to a highly dynamic regulation of the H-NOX domain. This competitive, bi-directional mode of interaction is also confirmed by monitoring cGMP generation in A7r5 vascular smooth muscle cells by these activators. We show that, surprisingly, heme's redox state impacts differently the bioactivity of these two structurally similar compounds. In all, by NMR-based and functional approaches we contribute unique experimental insight into the dynamic interaction of sGC activators with the H-NOX domain and its dependence on the heme redox status, with the ultimate goal to permit a better design of such therapeutically important molecules.

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“…In the review by Wittenborn et al the current view of full length sGC revealed by novel structures obtained by cryo-electron microscopy is summarized. The authors discussed the inactivated and activated states of the enzyme to describe the detailed biochemical mechanism and its function, which could be helpful to elucidate new sGC modulators for pharmacological treatments [ 6 ].…”

Section: Communicationmentioning

confidence: 99%