Spectroscopic and computational investigations of Cobalt(II) binding to the innate immune protein human calprotectin.

Human calprotectin (CP) is an innate immune protein that participates in the metal-withholding response to infection by sequestering essential metal nutrients from invading microbial pathogens. CP is comprised of S100A8 (α subunit, 10.8 kDa) and S100A9 (ß subunit, 13.2 kDa). Two transition-metal binding sites of CP form at the S100A8/S100A9 dimer interface. Site 1 is a His3Asp motif comprised of His83 and His87 from the S100A8 subunit and His20 and Asp30 from the S100A9 subunit. Site 2 is an unusual hexahistidine motif composed of S100A8 residues His17 and His27 and S100A9 residues His91, His95, His103, and His105. In the present study, the His3Asp and His6 sites of CP were further characterized by utilizing Co2+ as a spectroscopic probe. Magnetic circular dichroism spectroscopy was employed in conjunction with electron paramagnetic resonance spectroscopy and density functional theory computations to characterize the Co2+-bound S100A8(C42S)/S100A9(C3S) CP-Ser variant and six site variants that allowed the His3Asp and His6 sites to be further probed. Our results provide new insight into the metal-binding sites of CP-Ser and the effect of amino acid substitutions on the structure of site 2.

Sacrificial Cobalt-Carbon Bond Homolysis in Coenzyme B12 as a Cofactor Conservation Strategy.

A sophisticated intracellular trafficking pathway in humans is used to tailor vitamin B12 into its active cofactor forms, and to deliver it to two known B12-dependent enzymes. Herein, we report an unexpected strategy for cellular retention of B12, an essential and reactive cofactor. If methylmalonyl-CoA mutase is unavailable to accept the coenzyme B12 product of adenosyltransferase, the latter catalyzes homolytic scission of the cobalt-carbon bond in an unconventional reversal of the nucleophilic displacement reaction that was used to make it. The resulting homolysis product binds more tightly to adenosyltransferase than does coenzyme B12, facilitating cofactor retention. We have trapped, and characterized spectroscopically, an intermediate in which the cobalt-carbon bond is weakened prior to being broken. The physiological relevance of this sacrificial catalytic activity for cofactor retention is supported by the significantly lower coenzyme B12 concentration in patients with dysfunctional methylmalonyl-CoA mutase but normal adenosyltransferase activity.

Electronic Structure of Anilinopyridinate-Supported Ru25+ Paddlewheel Compounds.

The electronic structures of the diruthenium compounds Ru2(ap)4Cl (1, ap = 2-anilinopyridinate) and Ru2(ap)4OTf (2) were investigated with UV-vis, resonance Raman, and magnetic circular dichroism (MCD) spectroscopies; SQUID magnetometry; and density functional theory (DFT) calculations. Both compounds have quartet spin ground states with large axial zero-field splitting of ∼60 cm-1 that is characteristic of Ru25+ compounds having a (π*, δ*)3 electron configuration and a Ru-Ru bond order of ∼2.5. Two major visible absorption features are observed at ∼770 and 430 nm in the electronic spectra, the assignments of which have previously been ambiguous. Both bands have significant charge-transfer character with some contributions from d → d transitions. MCD spectra were measured to enable the identification of d → d transitions that are not easily observable by UV-vis spectroscopy. In this way, we are able to identify bands due to δ → δ* and δ → π* transitions at ∼16 100 and 11 200-12 300 cm-1, respectively, the latter band being sensitive to the π-donating character of the axial ligand. The Ru-Ru stretches are coupled with pyridine rocking motions and give rise to observed resonance Raman peaks at ∼350 and 420 cm-1, respectively.

Electronic, Magnetic, and Redox Properties and O2 Reactivity of Iron(II) and Nickel(II) o-Semiquinonate Complexes of a Tris(thioether) Ligand: Uncovering the Intradiol Cleaving Reactivity of an Iron(II) o-Semiquinonate Complex.

The iron(II) semiquinonate character within the iron(III) catecholate species has been proposed by numerous studies to account for the O2 reactivity of intradiol catechol dioxygenases, but a well-characterized iron(II) semiquinonate species that exhibits intradiol cleaving reactivity has not yet been reported. In this study, a detailed electronic structure description of the first iron(II) o-semiquinonate complex, [PhTttBu]Fe(phenSQ) [PhTttBu = phenyltris(tert-butylthiomethyl)borate; phenSQ = 9,10-phenanthrenesemiquinonate; Wang et al. Chem. Commun. 2014, 50, 5871-5873], was generated through a combination of electronic and Mössbauer spectroscopies, SQUID magnetometry, and density functional theory (DFT) calculations. [PhTttBu]Fe(phenSQ) reacts with O2 to generate an intradiol cleavage product, diphenic anhydride, in 16% yield. To assess the dependence of the intradiol reactivity on the identity of the metal ion, the nickel analogue, [PhTttBu]Ni(phenSQ), and its derivative, [PhTttBu]Ni(3,5-DBSQ) (3,5-DBSQ = 3,5-di-tert-butyl-1,2-semiquinonate), were prepared and characterized by X-ray crystallography, mass spectrometry, 1H NMR and electronic spectroscopies, and SQUID magnetometry. DFT calculations, evaluated on the basis of the experimental data, support the electronic structure descriptions of [PhTttBu]Ni(phenSQ) and [PhTttBu]Ni(3,5-DBSQ) as high-spin nickel(II) complexes with antiferromagnetically coupled semiquinonate ligands. Unlike its iron counterpart, [PhTttBu]Ni(phenSQ) decomposes slowly in an O2 atmosphere to generate 14% phenanthrenequinone with a negligible amount of diphenic anhydride. [PhTttBu]Ni(3,5-DBSQ) does not react with O2. This dramatic effect of the metal-ion identity supports the hypothesis that a metal(III) alkylperoxo species serves as an intermediate in the intradiol cleaving reactions. The redox properties of all three complexes were probed using cyclic voltammetry and differential pulse voltammetry, which indicate an inner-sphere electron-transfer mechanism for the formation of phenanthrenequinone. The lack of O2 reactivity of [PhTttBu]Ni(3,5-DBSQ) can be rationalized by the high redox potential of the metal-ligated 3,5-DBSQ/3,5-DBQ couple.

Dependence of surface-enhanced infrared absorption (SEIRA) enhancement and spectral quality on the choice of underlying substrate: a closer look at silver (Ag) films prepared by physical vapor deposition (PVD).

Silver (Ag) films of varying thickness were simultaneously deposited using physical vapor deposition (PVD) onto six infrared (IR) substrates (BaF(2), CaF(2), Ge, AMTIR, KRS-5, and ZnSe) in order to correlate the morphology of the deposited film with optimal SEIRA response and spectral band symmetry and quality. Significant differences were observed in the surface morphology of the deposited silver films, the degree of enhancement provided, and the spectral appearance of para-nitrobenzoic acid (PNBA) cast films for each silver-coated substrate. These differences were attributed to each substrate's chemical properties, which dictate the morphology of the Ag film and ultimately determine the spectral appearance of the adsorbed analyte and the magnitude of SEIRA enhancement. Routine SEIRA enhancement factors (EFs) for all substrates were between 5 and 150. For single-step Ag depositions, the following ranking identifies the greatest SEIRA enhancement factor and the maximum absorption of the 1345 cm(-1) spectral marker of PNBA at the optimal silver thickness for each substrate: BaF(2) (EF = 85 ± 19, 0.059 A, 10 nm Ag) > CaF(2) (EF = 75 ± 30, 0.052 A, 10 nm Ag) > Ge (EF = 45 ± 8, 0.019 A, 5 nm Ag) > AMTIR (EF = 38 ± 8, 0.024 A, 15 nm Ag) > KRS-5 (EF = 24 ± 1, 0.015 A, 12 nm Ag) > ZnSe (EF = 9 ± 5, 0.008 A, 8 nm Ag). A two-step deposition provides 59% larger EFs than single-step depositions of Ag on CaF(2). A maximum EF of 147 was calculated for a cast film of PNBA (surface coverage = 341 ng/cm(2)) on a 10 nm two-step Ag film on CaF(2) (0.102 A, 1345 cm(-1) symmetric NO(2) stretching band). The morphology of the two-step Ag film has smaller particles and greater particle density than the single-step Ag film.