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Heme-based globin-coupled oxygen sensors: linking oxygen binding to functional regulation of diguanylate cyclase, histidine kinase, and methyl-accepting chemotaxis.

ABSTRACT: An emerging class of novel heme-based oxygen sensors containing a globin fold binds and senses environmental O2 via a heme iron complex. Structure-function relationships of oxygen sensors containing a heme-bound globin fold are different from those containing heme-bound PAS and GAF folds. It is thus worth reconsidering from an evolutionary perspective how heme-bound proteins with a globin fold similar to that of hemoglobin and myoglobin could act as O2 sensors. Here, we summarize the molecular mechanisms of heme-based oxygen sensors containing a globin fold in an effort to shed light on the O2-sensing properties and O2-stimulated catalytic enhancement observed for these proteins.

SUBMITTER: Martinkova M

PROVIDER: S-EPMC3784688 | biostudies-literature | 2013 Sep

REPOSITORIES: biostudies-literature

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Heme-based globin-coupled oxygen sensors: linking oxygen binding to functional regulation of diguanylate cyclase, histidine kinase, and methyl-accepting chemotaxis.

Martínková Markéta M Kitanishi Kenichi K Shimizu Toru T

The Journal of biological chemistry 20130808 39

An emerging class of novel heme-based oxygen sensors containing a globin fold binds and senses environmental O2 via a heme iron complex. Structure-function relationships of oxygen sensors containing a heme-bound globin fold are different from those containing heme-bound PAS and GAF folds. It is thus worth reconsidering from an evolutionary perspective how heme-bound proteins with a globin fold similar to that of hemoglobin and myoglobin could act as O2 sensors. Here, we summarize the molecular m ...[more]

PMID: 23928310

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Project description:The periplasmic sensor domains GSU0582 and GSU0935 are part of methyl accepting chemotaxis proteins in the bacterium Geobacter sulfurreducens. Both contain one c-type heme group and their crystal structures revealed that these domains form swapped dimers with a PAS fold formed from the two protein chains. The swapped dimerization of these sensors is related to the mechanism of signal transduction and the formation of the swapped dimer involves significant folding changes and conformational rearrangements within each monomeric component. However, the structural changes occurring during this process are poorly understood and lack a mechanistic framework. To address this issue, we have studied the folding and stability properties of two distinct heme-sensor PAS domains, using biophysical spectroscopies. We observed substantial differences in the thermodynamic stability (?G?=?14.6 kJ.mol(-1) for GSU0935 and ?G?=?26.3 kJ.mol(-1) for GSU0582), and demonstrated that the heme moiety undergoes conformational changes that match those occurring at the global protein structure. This indicates that sensing by the heme cofactor induces conformational changes that rapidly propagate to the protein structure, an effect which is directly linked to the signal transduction mechanism. Interestingly, the two analyzed proteins have distinct levels of intrinsic disorder (25% for GSU0935 and 13% for GSU0582), which correlate with conformational stability differences. This provides evidence that the sensing threshold and intensity of the propagated allosteric effect is linked to the stability of the PAS-fold, as this property modulates domain swapping and dimerization. Analysis of the PAS-domain shows that disorder segments are found either at the hinge region that controls helix motions or in connecting segments of the ?-sheet interface. The latter is known to be widely involved in both intra- and intermolecular interactions, supporting the view that it's folding and stability are at the basis of the specificity and regulation of many types of PAS-containing signaling proteins.

Project description:SignificanceLigand selectivity for dioxygen (O(2)), carbon monoxide (CO), and nitric oxide (NO) is critical for signal transduction and is tailored specifically for each heme-protein sensor. Key NO sensors, such as soluble guanylyl cyclase (sGC), specifically recognized low levels of NO and achieve a total O(2) exclusion. Several mechanisms have been proposed to explain the O(2) insensitivity, including lack of a hydrogen bond donor and negative electrostatic fields to selectively destabilize bound O(2), distal steric hindrance of all bound ligands to the heme iron, and restriction of in-plane movements of the iron atom.Recent advancesCrystallographic structures of the gas sensors, Thermoanaerobacter tengcongensis heme-nitric oxide/oxygen-binding domain (Tt H-NOX(1)) or Nostoc puntiforme (Ns) H-NOX, and measurements of O(2) binding to site-specific mutants of Tt H-NOX and the truncated β subunit of sGC suggest the need for a H-bonding donor to facilitate O(2) binding.Critical issuesHowever, the O(2) insensitivity of full length sGC with a site-specific replacement of isoleucine by a tyrosine on residue 145 and the very slow autooxidation of Ns H-NOX and cytochrome c' suggest that more complex mechanisms have evolved to exclude O(2) but retain high affinity NO binding. A combined graphical analysis of ligand binding data for libraries of heme sensors, globins, and model heme shows that the NO sensors dramatically inhibit the formation of six-coordinated NO, CO, and O(2) complexes by direct distal steric hindrance (cyt c'), proximal constraints of in-plane iron movement (sGC), or combinations of both following a sliding scale rule. High affinity NO binding in H-NOX proteins is achieved by multiple NO binding steps that produce a high affinity five-coordinate NO complex, a mechanism that also prevents NO dioxygenation.Future directionsKnowledge advanced by further extensive test of this "sliding scale rule" hypothesis should be valuable in guiding novel designs for heme based sensors.

Project description: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.

Dataset Information

Heme-based globin-coupled oxygen sensors: linking oxygen binding to functional regulation of diguanylate cyclase, histidine kinase, and methyl-accepting chemotaxis.

Publications

Heme-based globin-coupled oxygen sensors: linking oxygen binding to functional regulation of diguanylate cyclase, histidine kinase, and methyl-accepting chemotaxis.

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