NO Adducts Research Articles

Photolability of NO in ruthenium nitrosyls with pentadentate ligand induces exceptional cytotoxicity towards VCaP, 22Rv1 and A549 cancer cells under therapeutic condition

Pentadentate electron rich MePBITA ligand in [RuII(MePBITA)(NO)]n+ (n = 3, 2 and MePBITA = 1-(6-(1-methyl-1H-benzo[d]imidazol-2-yl)pyridin-2-yl)-N,N-bis(pyridin-2-ylmethyl)methanamine) permits the isolation of both the redox states of nitrosyls with Enemark–Feltham notation {RuNO}6 and {RuNO}7. The nitrosyl derivative [RuII(MePBITA)(NO)](ClO4)3: [4](ClO4)3 was synthesized by stepwise synthetic manner from the chloro precursor [RuII(MePBITA)(Cl)](PF6): [1](PF6), via the acetonitrile derivative [RuII(MePBITA)(CH3CN)](PF6)2: [2](PF6)2 followed by nitro complex [RuII(MePBITA)(NO2)](PF6): [3](PF6). All the complexes were fully characterized by different analytical and spectroscopic techniques. Single crystal X-ray structures of the complexes [1](PF6), [2](PF6)2, [3](PF6), and [4](ClO4)3 were profitably determined for understanding the molecular integrity. Ru−NO stretching frequency observed at 1931 cm−1 for [4](ClO4)3 suggests a moderately electrophilic character of NO. The huge shift in νNO frequency, Δν (solid) = 325 cm−1 was observed by reducing [4](ClO4)3 to [4](ClO4)2. The conversion of [3]+ from [4]3+ was examined both electrochemically and spectrophotometrically with the addition of 0.5 M NaOH solution. Rate constants of the first order photorelease (kNO) have been found to be 8.99 × 10−3 min−1; half-life (t1/2) = 77 min and 3.84 × 10−2 min−1; half-life (t1/2) = 18 min for [4]3+ and [4]2+, respectively with visible Xenon light (200 W) source. The photo liberated NO has been scavenged by biologically relevant target protein reduced myoglobin as Mb−NO adduct. Photoactivation of [4]3+ and [4]2+ by visible light induces significant cytotoxicity in prostate cancer cell lines; VCaP (IC50 29.74 and 4.42 µM) and 22Rv1 (IC50 29.96 and 6.88 µM), and lung cancer cell line; A549 (IC50 2.24 and 0.12 µM). Collectively our results pave the way for the development of metallodrugs as potential therapeutics for a variety of cancers. Additionally, our results also demonstrate how ligand modification could enhance the photolability of metal nitrosyl, adding a new dimension for future efficient photoactive metal nitrosyl design.

Insights into the Observed trans-Bond Length Variations upon NO Binding to Ferric and Ferrous Porphyrins with Neutral Axial Ligands.

NO is well-known for its trans effect. NO binding to ferrous hemes of the form (por)Fe(L) (L = neutral N-based ligand) to give the {FeNO}7 (por)Fe(NO)(L) product results in a lengthening of the axial trans Fe–L bond. In contrast, NO binding to the ferric center in [(por)Fe(L)]+ to give the {FeNO}6 [(por)Fe(NO)(L)]+ product results in a shortening of the trans Fe–L bond. NO binding to both ferrous and ferric centers involves the lowering of their spin states. Density functional theory (DFT) calculations were used to probe the experimentally observed trans-bond shortening in some NO adducts of ferric porphyrins. We show that the strong σ antibonding interaction of dz2 and the axial (L) ligand p orbitals present in the Fe(II) systems is absent in the Fe(III) systems, as it is now in an unoccupied orbital. This feature, combined with a lowering of spin state upon NO binding, provides a rationale for the observed net trans-bond shortening in the {FeNO}6 but not the {FeNO}7 derivatives.

Oxygen-mediated oxidation of ferrous nitrosylated nitrobindins

The O2-mediated oxidation of all-β-barrel ferrous nitrosylated nitrobindin from Arabidopsis thaliana (At-Nb(II)-NO), Mycobacterium tuberculosis (Mt-Nb(II)-NO), and Homo sapiens (Hs-Nb(II)-NO) to ferric derivative (At-Nb(III), Mt-Nb(III), and Hs-Nb(III), respectively) has been investigated at pH 7.0 and 20.0 °C. Unlike ferrous nitrosylated horse myoglobin, human serum heme-albumin and human hemoglobin, the process in Nb(II)-NO is mono-exponential and linearly dependent on the O2 concentration, displaying a bimolecular behavior, characterized by kon = (6.3 ± 0.8) × 103 M−1 s−1, (1.4 ± 0.2) × 103 M−1 s−1, and (3.9 ± 0.5) × 103 M−1 s−1 for At-Nb(II)-NO, Mt-Nb(II)-NO, and Hs-Nb(II)-NO, respectively. No intermediate is detected, indicating that the O2 reaction with Nb(II)-NO is the rate-limiting step and that the subsequent conversion of the heme-Fe(III)-N(O)OO− species (i.e., N-bound peroxynitrite to heme-Fe(III)) to heme-Fe(III) and NO3− is much faster. A similar mechanism can be invoked for ferrous nitrosylated human neuroglobin and rabbit hemopexin, in which the heme-Fe(III)-N(O)OO− species is formed as well, although the rate-limiting step seems represented by the reshaping of the six-coordinated heme-Fe(III) complex. Although At-Nb(II)-NO and Mt-Nb(II)-NO are partially (while Hs-Nb(II)-NO is almost completely) penta-coordinated, density functional theory (DFT) calculations rule out that the cleavage of the proximal heme-Fe-His bond in Nb(II)-NO is responsible for the more stable heme-Fe(III)-N(O)OO− species. Moreover, the oxidation of the penta-coordinated heme-Fe(II)-NO adduct does not depend on O2 binding at the proximal side of the metal center. These features may instead reflect the peculiarity of Nb folding and of the heme environment, with a reduced steric constraint for the formation of the heme-Fe(III)-N(O)OO− complex.

Melatonin, Its Metabolites and Their Interference with Reactive Nitrogen Compounds.

Melatonin and several of its metabolites are interfering with reactive nitrogen. With the notion of prevailing melatonin formation in tissues that exceeds by far the quantities in blood, metabolites come into focus that are poorly found in the circulation. Apart from their antioxidant actions, both melatonin and N1-acetyl-5-methoxykynuramine (AMK) downregulate inducible and inhibit neuronal NO synthases, and additionally scavenge NO. However, the NO adduct of melatonin redonates NO, whereas AMK forms with NO a stable product. Many other melatonin metabolites formed in oxidative processes also contain nitrosylatable sites. Moreover, AMK readily scavenges products of the CO2-adduct of peroxynitrite such as carbonate radicals and NO2. Protein AMKylation seems to be involved in protective actions.

High‐spin square planar iron(II) alkali metal siloxide complexes – influence of the alkali metal and reactivity towards O2 and NO

AbstractComplexes [L2Fe][Li(DME)]2, 1(DME), {[L2Fe][Na2(DME)3]}∞, 2(DME) and [L2Fe][K(DME)2]2, 3(DME) were synthesized by deprotonation of LH2 (LH2=O(SiPh2OH)2) with the respective alkali metal tert‐butoxides followed by recrystallization from DME. It turned out that upon crossing over from Li+ via Na+ to K+ counterions the structures of the high‐spin iron(II) complexes are increasingly distorted from a square planar towards a tetrahedral structure so that 3(DME) represents a borderline case, as indicated by the τ‐values. The distortions are also reflected in the Mössbauer spectra through the quadrupole splittings. The compounds behave inert in contact with O atom transfer reagents but react rapidly with dioxygen. The reaction rates are too high to be determined even by stopped‐flow measurements quantitatively, but qualitatively it emerged that the rates increase from Li to Na to K. Using NO as an O2 surrogate an NO adduct with an S = 3/2 ground state was isolated where NO is coordinated in an end‐on binding mode, formally as a NO− ligand, with a significantly weakened NO bond.

Structure and mechanistic relevance of Ni2+–NO adduct in model HC SCR reaction over NiZSM-5 catalyst – Insights from standard and correlation EPR and IR spectroscopic studies corroborated by molecular modeling

Mechanistic aspects of model selective catalytic reduction (SCR) of NO with C2H4 over Ni/ZSM-5 zeolite were investigated by means of advanced correlation EPR/HYSCORE and IR time-resolved Rapid-Scan 2D COS techniques combined with DFT/CASSCF calculations. It was shown that two types of the active centers, bare isolated Ni2+ and dual oxo Ni2+–O2––Ni2+, are involved in this reaction, acting in different ways. NO interaction with the former center results in a Ni2+–NOδ+ moiety, the unique nature of which was ascertained by HYSCORE studies, and accounted for by the spin-polarized DFT/CASSCF calculations in detail. The electrophilic nature of the bound NOδ+ moiety is crucial for its successful insertion into the >C=C< bond of ethylene. This process involves formation of isolated nickel adducts with the co-ligated C2H4 and NO molecules, and the resultant cyanide and isocyanate intermediates are next hydrolyzed into ammonia or oxidized by NOx. The dual nickel-oxo centers provide the active sites for development of nitrate/nitro species and next mixed-ligand nitrate/nitro-ammonia adducts. Selective catalytic reduction results from the NOx/cyanide and nitrate/nitro-ammonia routes, which both are featured by reversal of the partial charge on the nitrogen atom from positive in Nδ+O into negative in –CNδ–/–Nδ–CO or Nδ–H3, giving rise to the key nitrogen charge comproportionation step. In both pathways, the nickel(II) mononitrosyls play a pivotal function as the primary source of the N–containing intermediates (–CN/–NCO/NH3). It acts also as a species hindering the undesired oxidation of C2H4 by dioxygen.

Interaction of nitric oxide with the (1 0 0) surface of cobalt spinel nanocubes – A comprehensive DFT, atomistic thermodynamic, IR and TPD account

Periodic spin-unrestricted DFT + U/PW91 calculations, atomistic thermodynamic modeling and IR and TPD measurements were used to study structure and stability of various NO adspecies on the (1 0 0) surface of Co3O4 nanocubes. A large variety of monodentate (η1), bridging bidentate (µ-η2) and bridging monodentate (µ-η1:η1) adducts were identified. The most stable were mononitrosyl (η1-ON-CoT) and dinitrosyl (η1-(ON)2-CoT) complexes of tetrahedral CoT centers, whereas the NO adducts of the octahedral CoO sites (η1-ON-CoO) were less favorable. The DOS structure, atomic partial charge and magnetization analysis revealed that their electronic structures can be epitomized as η1-ON-CoT↑↑ and η1-ON↑CoO↓↑. Thermodynamic ΘNO = f(pNO,T) diagrams were constructed and used for interpretation of a three-step NO desorption corresponding to decay of η1-ON-CoO at 60 ÷ 80 °C, decomposition of η1-(ON)2-CoT at 210 °C, and decay of η1-ON-CoT at 380 °C. The stability and spectroscopic signatures of these adspecies were ascertained in IR and NO-TPD measurements. The 1940 cm−1 band corresponds to chemisorption of NO on CoO sites, whereas for the CoT centers the mononitrosyl species appear at 1850 cm−1 and the dinitrosyls at 1895 and 1813 cm−1. The TPD profiles confirm the three NO desorption peaks in the temperature windows, predicted by the DFT and thermodynamic modelling.

Fe Speciation in Iron Modified Natural Zeolites as Sustainable Environmental Catalysts

Natural purified mordenite from Palmarito de Cauto (ZP) deposit, Cuba, was subjected to a hydrothermal ion exchange process in acid medium with Fe2+ or Fe3+ salts (Fe2+ZP and Fe3+ZP). The set of samples was characterized regarding their textural properties, morphology, and crystallinity, and tested in the NO reduction with CO/C3H6. Infrared spectroscopy coupled with NO as a probe molecule was used to give a qualitative description of the Fe species’ nature and distribution. The exchange process caused an increase in the iron loading of the samples and a redistribution, resulting in more dispersed Fe2+ and Fe3+ species. When contacted with the NO probe, Fe2+ZP showed the highest intensity of nitrosyl bands, assigned to NO adducts on isolated/highly dispersed Fe2+/Fe3+ extra-framework sites and FexOy clusters. This sample is also characterized by the highest NO sorption capacity and activity in NO reduction. Fe3+ZP showed a higher intensity of nitrosonium (NO+) species, without a correlation to NO storage and conversion, pointing to the reactivity of small FexOy aggregates in providing oxygen atoms for the NO to NO+ reaction. The same sites are proposed to be responsible for the higher production of CO2 observed on this sample, and thus to be detrimental to the activity in NO SCR.

Non-heme High-Spin {FeNO}6-8 Complexes: One Ligand Platform Can Do It All.

Heme and non-heme iron-nitrosyl complexes are important intermediates in biology. While there are numerous examples of low-spin heme iron-nitrosyl complexes in different oxidation states, much less is known about high-spin (hs) non-heme iron-nitrosyls in oxidation states other than the formally ferrous NO adducts ({FeNO}7 in the Enemark-Feltham notation). In this study, we present a complete series of hs-{FeNO}6-8 complexes using the TMG3tren coligand. Redox transformations from the hs-{FeNO}7 complex [Fe(TMG3tren)(NO)]2+ to its {FeNO}6 and {FeNO}8 analogs do not alter the coordination environment of the iron center, allowing for detailed comparisons between these species. Here, we present new MCD, NRVS, XANES/EXAFS, and Mössbauer data, demonstrating that these redox transformations are metal based, which allows us to access hs-Fe(II)-NO-, Fe(III)-NO-, and Fe(IV)-NO- complexes. Vibrational data, analyzed by NCA, directly quantify changes in Fe-NO bonding along this series. Optical data allow for the identification of a "spectator" charge-transfer transition that, together with Mössbauer and XAS data, directly monitors the electronic changes of the Fe center. Using EXAFS, we are also able to provide structural data for all complexes. The magnetic properties of the complexes are further analyzed (from magnetic Mössbauer). The properties of our hs-{FeNO}6-8 complexes are then contrasted to corresponding, low-spin iron-nitrosyl complexes where redox transformations are generally NO centered. The hs-{FeNO}8 complex can further be protonated by weak acids, and the product of this reaction is characterized. Taken together, these results provide unprecedented insight into the properties of biologically relevant non-heme iron-nitrosyl complexes in three relevant oxidation states.

Detection of Copper Bisguanidine NO Adducts by UV‐vis Spectroscopy and a SuperFocus Mixer

AbstractThe reaction of Cu(I) bisguanidine complexes with nitric oxide and the formation of intermediate species were monitored via UV‐vis spectroscopy at low temperature, with the occurrence of characteristic absorption bands. The origin of the emerging species and their character were substantiated by electron paramagnetic resonance (EPR) measurements and density functional theory (DFT) studies showing a delocalized {CuNO}11 radical species. Furthermore, this system was transferred to the SuperFocus mixer setup, which allows rapid mixing and the determination of decay constants at ambient temperatures of the thermally sensitive species. However, these experiments demonstrated the limits of these systems, such as the NO saturation in organic solvents and a preferably precise temperature control within the SuperFocus mixer, which should be addressed in the future.

Resonance Raman in Vitro Detection and Differentiation of the Nitrite-Induced Hemoglobin Adducts in Functional Human Red Blood Cells.

This work presents in vitro studies of the functional, isolated human red blood cells (RBCs) treated with various concentrations of Na14NO2 and Na15NO2 with the use of resonance Raman spectroscopy (RRS) at two different laser excitations supported by absorption spectrophotometry (UV-vis). The products of the reaction between oxyhemoglobin (oxyHb) in isolated RBCs with NaNO2 were analyzed and identified in situ. The metHb-H2O was found to be the major product of this reaction; however, additional adducts were also clearly observed. Vibrational analysis allowed identification of various Hb3+NO2 species (Fe3+-O-N=O with O-binding mode of nitrite ion to the Fe3+ core and nitrovinyl adducts with 2-vinyl nitration favored over 4-vinyl nitration) as well as the Fe3+-NO adduct. In addition, we were able to visualize in situ the Hb-NO2 species inside functional RBCs with the use of Raman imaging.

CO and NO bind to Fe(II) DiGeorge critical region 8 heme but do not restore primary microRNA processing activity.

The RNA-binding heme protein DiGeorge critical region 8 (DGCR8) and its ribonuclease partner Drosha cleave primary transcripts of microRNA (pri-miRNA) as part of the canonical microRNA (miRNA) processing pathway. Previous studies show that bis-cysteine thiolate-coordinated Fe(III) DGCR8 supports pri-miRNA processing activity, while Fe(II) DGCR8 does not. In this study, we further characterized Fe(II) DGCR8 and tested whether CO or NO might bind and restore pri-miRNA processing activity to the reduced protein. Fe(II) DGCR8 RNA-binding heme domain (Rhed) undergoes a pH-dependent transition from 6-coordinate to 5-coordinate, due to protonation and loss of a lysine ligand; the ligand bound throughout the pH change is a histidine. Fe(II) Rhed binds CO and NO from 6- and 5-coordinate states, forming common CO and NO adducts at all pHs. Fe(II)-CO Rhed is 6-coordinate, low-spin, and pH insensitive with the histidine ligand retained, suggesting that the protonatable lysine ligand has been replaced by CO. Fe(II)-NO Rhed is 5-coordinate and pH insensitive. Fe(II)-NO also forms slowly upon reaction of Fe(III) Rhed with excess NO via a stepwise process. Heme reduction by NO is rate-limiting, and the rate would be negligible at physiological NO concentrations. Importantly, in vitro pri-miRNA processing assays show that both CO- and NO-bound DGCR8 species are inactive. Fe(II), Fe(II)-CO, and Fe(II)-NO Rhed do not bear either of the cysteine ligands found in the Fe(III) state. These data support a model in which the bis-cysteine thiolate ligand environment of Fe(III) DGCR8 is necessary for establishing proper pri-miRNA binding and enabling processing activity.

Structural and Spectroscopic Characterization of a High‐Spin {FeNO}6 Complex with an Iron(IV)−NO− Electronic Structure

AbstractAlthough the interaction of low‐spin ferric complexes with nitric oxide has been well studied, examples of stable high‐spin ferric nitrosyls (such as those that could be expected to form at typical non‐heme iron sites in biology) are extremely rare. Using the TMG3tren co‐ligand, we have prepared a high‐spin ferric NO adduct ({FeNO}6 complex) via electrochemical or chemical oxidation of the corresponding high‐spin ferrous NO {FeNO}7 complex. The {FeNO}6 compound is characterized by UV/Visible and IR spectroelectrochemistry, Mössbauer and NMR spectroscopy, X‐ray crystallography, and DFT calculations. The data show that its electronic structure is best described as a high‐spin iron(IV) center bound to a triplet NO− ligand with a very covalent iron−NO bond. This finding demonstrates that this high‐spin iron nitrosyl compound undergoes iron‐centered redox chemistry, leading to fundamentally different properties than corresponding low‐spin compounds, which undergo NO‐centered redox transformations.

Structural and Spectroscopic Characterization of a High-Spin {FeNO}(6) Complex with an Iron(IV)-NO(-) Electronic Structure.

Although the interaction of low-spin ferric complexes with nitric oxide has been well studied, examples of stable high-spin ferric nitrosyls (such as those that could be expected to form at typical non-heme iron sites in biology) are extremely rare. Using the TMG3 tren co-ligand, we have prepared a high-spin ferric NO adduct ({FeNO}(6) complex) via electrochemical or chemical oxidation of the corresponding high-spin ferrous NO {FeNO}(7) complex. The {FeNO}(6) compound is characterized by UV/Visible and IR spectroelectrochemistry, Mössbauer and NMR spectroscopy, X-ray crystallography, and DFT calculations. The data show that its electronic structure is best described as a high-spin iron(IV) center bound to a triplet NO(-) ligand with a very covalent iron-NO bond. This finding demonstrates that this high-spin iron nitrosyl compound undergoes iron-centered redox chemistry, leading to fundamentally different properties than corresponding low-spin compounds, which undergo NO-centered redox transformations.

Nitric Oxide Catalysis of Diazene E/Z Isomerization.

Nitric oxide is an efficient catalyst for the cis-trans (E/Z) isomerization of diazenes. We compare the effect of room temperature solutions bearing low concentrations of nitric oxide, nitrogen dioxide, or oxygen on the rate of cis-trans isomerization, CTI, of the alkene bond in stilbene and on the azo double bond in azobenzene, as well as in four azo derivatives as measured by UV-vis spectroscopy. These rate enhancements can be as large as 3 orders of magnitude for azobenzene in solution. A mechanism is proposed where catalysis is promoted by the interaction of the nitric oxide with the diazene nitrogen lone pairs. Density functional theory, B3LYP/6-311++g** suggests that the binding of NO to the diazene should be weak and reversible but that its NO adduct has an E/Z isomerization barrier of 7.5 kcal/mol.

Hydrogen bonding of the dissociated histidine ligand is not required for formation of a proximal NO adduct in cytochrome c'.

Cytochromes c', that occur in methanotrophic, denitrifying and photosynthetic bacteria, form unusual proximal penta-coordinate NO complexes via a hexa-coordinate distal NO intermediate. Their NO binding properties are similar to those of the eukaryotic NO sensor, soluble guanylate cyclase, for which they provide a valuable structural model. Previous studies suggested that hydrogen bonding between the displaced proximal histidine (His120) ligand (following its dissociation from heme due to trans effects from the distally bound NO) and a conserved aspartate residue (Asp121) could play a key role in allowing proximal NO binding to occur. We have characterized three variants of Alcaligenes xylosoxidans cytochrome c' (AXCP) where Asp121 has been replaced by Ala, Ile and Gln, respectively. In all variants, hydrogen bonding between residue 121 and His120 is abolished yet 5-coordinate proximal NO species are still formed. Our data therefore demonstrate that the His120-Asp121 bond is not essential for proximal NO binding although it likely provides an energy minimum for the displaced His ligand. All variants have altered proximal pocket structure relative to native AXCP.

Solid-state EPR strategies for the structural characterization of paramagnetic NO adducts of frustrated Lewis pairs (FLPs).

Anisotropic interactions present in three new nitroxide radicals prepared by N,N addition of NO to various borane-phosphane frustrated Lewis pairs (FLPs) have been characterized by continuous-wave (cw) and pulsed X-band EPR spectroscopies in solid FLP-hydroxylamine matrices at 100 K. Anisotropic g-tensor values and (11)B, (14)N, and (31)P hyperfine coupling tensor components have been extracted from continuous-wave lineshape analyses, electron spin echo envelope modulation (ESEEM), and hyperfine sublevel correlation spectroscopy (HYSCORE) experiments with the help of computer simulation techniques. Suitable fitting constraints are developed on the basis of density functional theory (DFT) calculations. These calculations reveal that different from the situation in standard nitroxide radicals (TEMPO), the g-tensors are non-coincident with any of the nuclear hyperfine interaction tensors. The determination of these interaction parameters turns out to be successful, as the cw- and pulse EPR experiments are highly complementary in informational content. While the continuous-wave lineshape is largely influenced by the anisotropic hyperfine coupling to (14)N and (31)P, the ESEEM and HYSCORE spectra contain important information about the (11)B hyperfine coupling and nuclear electric quadrupolar interaction. The set of cw- and pulsed EPR experiments, with fitting constraints developed by DFT calculations, defines an efficient strategy for the structural analysis of paramagnetic FLP adducts.

Nitrosyl–heme and anion–arene complexes: structure, reactivity and spectroscopy

Abstract Two topics are selected and illustrated to exemplify (i) a biological and (ii) an organic ionic intermediate. The reactivity behavior of NO adducts with ferric and ferrous hemes has shown remarkable similarities when examined in the gas phase, demonstrating that the largely different NO affinity displayed in solution and in biological media is due to the different coordination environment. In fact, ferrous hemes present a vacant or highly labile axial coordination site, prone to readily bind NO. The vibrational signatures of the NO ligand have also been probed in vacuo for the first time in the nitrosyl complexes deriving from ferrous and ferric hemes under strictly comparable five-coordination at the metal center. Negatively charged σ-adducts, from the association of anions with 1,3,5-trinitrobenzene, an exemplary π-electron-deficient arene, have been probed by IRMPD spectroscopy and found to display variable binding motifs from a strongly covalent σ-adduct (Meisenheimer complex) to a weakly covalent σ-complex, depending on the anion basicity.

Conformational control of the binding of diatomic gases to cytochrome c'.

The cytochromes c' (CYTcp) are found in denitrifying, methanotrophic and photosynthetic bacteria. These proteins are able to form stable adducts with CO and NO but not with O2. The binding of NO to CYTcp currently provides the best structural model for the NO activation mechanism of soluble guanylate cyclase. Ligand binding in CYTcps has been shown to be highly dependent on residues in both the proximal and distal heme pockets. Group 1 CYTcps typically have a phenylalanine residue positioned close to the distal face of heme, while for group 2, this residue is typically leucine. We have structurally, spectroscopically and kinetically characterised the CYTcp from Shewanella frigidimarina (SFCP), a protein that has a distal phenylalanine residue and a lysine in the proximal pocket in place of the more common arginine. Each monomer of the SFCP dimer folds as a 4-alpha-helical bundle in a similar manner to CYTcps previously characterised. SFCP exhibits biphasic binding kinetics for both NO and CO as a result of the high level of steric hindrance from the aromatic side chain of residue Phe 16. The binding of distal ligands is thus controlled by the conformation of the phenylalanine ring. Only a proximal 5-coordinate NO adduct, confirmed by structural data, is observed with no detectable hexacoordinate distal NO adduct.

Gradual release of strongly bound nitric oxide from Fe₂(NO)₂(dobdc).

An iron(II)-based metal-organic framework featuring coordinatively unsaturated redox-active metal cation sites, Fe2(dobdc) (dobdc(4-) = 2,5-dioxido-1,4-benzenedicarboxylate), is shown to strongly bind nitric oxide at 298 K. Adsorption isotherms indicate an adsorption capacity greater than 16 wt %, corresponding to the adsorption of one NO molecule per iron center. Infrared, UV-vis, and Mössbauer spectroscopies, together with magnetic susceptibility data, confirm the strong binding is a result of electron transfer from the Fe(II) sites to form Fe(III)-NO(-) adducts. Consistent with these results, powder neutron diffraction experiments indicate that NO is bound to the iron centers of the framework with an Fe-NO separation of 1.77(1) Å and an Fe-N-O angle of 150.9(5)°. The nitric oxide-containing material, Fe2(NO)2(dobdc), steadily releases bound NO under humid conditions over the course of more than 10 days, suggesting it, and potential future iron(II)-based metal-organic frameworks, are good candidates for certain biomedical applications.