Intermediates involved in the reduction of SO2: insight into the mechanism of sulfite reductases.

Sulfite reductases (SiRs) catalyze the reduction of SO32- to H2S in biosynthetic sulfur assimilation and dissimilation of sulfate. The mechanism of the 6e-/6H+ reduction of SO32- at the siroheme cofactor is debated, and proposed intermediates involved in this 6e- reduction are yet to be spectroscopically characterized. The reaction of SO2 with a ferrous iron porphyrin is investigated, and two intermediates are trapped and characterized: an initial Fe(III)-SO22- species, which undergoes proton-assisted S-O bond cleavage to form an Fe(III)-SO species. These species are characterized using a combination of resonance Raman (with 34S-labelled SO2), EPR and DFT calculations. Results obtained help reconcile the different proposed mechanisms for the SiRs.

Iron Dioxygen Adduct Formed during Electrochemical Oxygen Reduction by Iron Porphyrins Shows Catalytic Heme Dioxygenase Reactivity.

Heme dioxygenases oxidize the indole ring of tryptophan to kynurenine which is the first step in the biosynthesis of several important biomolecules like NAD, xanthurenic acid, and picolinic acid. A ferrous heme dioxygen adduct (or FeIII-O2•-) is the oxidant, and both the atoms of O2 are inserted in the product and its catalytic function has been difficult to emulate as it is complicated by competing rapid reactions like auto-oxidation and/or formation of the µ-oxo dimer. In situ resonance Raman spectroscopy technique, SERRS-RDE, is used to probe the species accumulated during electrochemical ORR catalyzed by site-isolated imidazole-bound iron porphyrin installed on a self-assembled monolayer covered electrode. These in situ SERRS-RDE data using labeled O2 show that indeed a FeIII-O2•- species accumulate on the electrode during ORR between -0.05 and -0.30 V versus Ag/AgCl (satd. KCl) and is reduced by proton coupled electron transfer to a FeIII-OOH species which, on the other hand, builds up on the electrode between -0.20 and -0.40 V versus Ag/AgCl (satd. KCl). This FeIII-OOH species then gives way to a FeIV═O species, which accumulates at -0.50 V versus Ag/AgCl (satd. KCl). When 2,3-dimethylindole is present in the solution and the applied potential is held in the range where FeIII-O2•- species accumulate, it gets oxidized to N-(2-acetylphenyl)acetamide retaining both the oxygens from O2 mimicking the reaction of heme dioxygenases. Turnover numbers more than 104 are recorded, establishing this imidazole-bound ferrous porphyrin as a functional model of heme dioxygenases.

Silver nanostructure-modified graphite electrode for in-operando SERRS investigation of iron porphyrins during high-potential electrocatalysis.

In-operando spectroscopic observation of the intermediates formed during various electrocatalytic oxidation and reduction reactions is crucial to propose the mechanism of the corresponding reaction. Surface-enhanced resonance Raman spectroscopy coupled to rotating disk electrochemistry (SERRS-RDE), developed about a decade ago, proved to be an excellent spectroscopic tool to investigate the mechanism of heterogeneous oxygen reduction reaction (ORR) catalyzed by synthetic iron porphyrin complexes under steady-state conditions in water. The information about the formation of the intermediates accumulated during the course of the reaction at the electrode interface helped to develop better ORR catalysts with second sphere residues in the porphyrin rings. To date, the application of this SERRS-RDE setup is limited to ORR only because the thiol self-assembled monolayer (SAM)-modified Ag electrode, used as the working electrode in these experiments, suffers from stability issues at more cathodic and anodic potential, where H2O oxidation, CO2 reduction, and H+ reduction reactions occur. The current investigation shows the development of a second-generation SERRS-RDE setup consisting of an Ag nanostructure (AgNS)-modified graphite electrode as the working electrode. These electrodes show higher stability (compared to the conventional thiol SAM-modified Ag electrode) upon exposure to very high cathodic and anodic potential with a good signal-to-noise ratio in the Raman spectra. The behavior of this modified electrode toward ORR is found to be the same as the SAM-modified Ag electrode, and the same ORR intermediates are observed during electrochemical ORR. At higher cathodic potential, the signatures of Fe(0) porphyrin, an important intermediate in H+ and CO2 reduction reactions, was observed at the electrode-water interface.

Second-Sphere Hydrogen-Bond Donors and Acceptors Affect the Rate and Selectivity of Electrochemical Oxygen Reduction by Iron Porphyrins Differently.

The factors that control the rate and selectivity of 4e-/4H+ O2 reduction are important for efficient energy transformation as well as for understanding the terminal step of respiration in aerobic organisms. Inspired by the design of naturally occurring enzymes which are efficient catalysts for O2 and H2O2 reduction, several artificial systems have been generated where different second-sphere residues have been installed to enhance the rate and efficiency of the 4e-/4H+ O2 reduction. These include hydrogen-bonding residues like amines, carboxylates, ethers, amides, phenols, etc. In some cases, improvements in the catalysis were recorded, whereas in some cases improvements were marginal or nonexistent. In this work, we use an iron porphyrin complex with pendant 1,10-phenanthroline residues which show a pH-dependent variation of the rate of the electrochemical O2 reduction reaction (ORR) over 2 orders of magnitude. In-situ surface-enhanced resonance Raman spectroscopy reveals the presence of different intermediates at different pH's reflecting different rate-determining steps at different pH's. These data in conjunction with density functional theory calculations reveal that when the distal 1,10-phenanthroline is neutral it acts as a hydrogen-bond acceptor which stabilizes H2O (product) binding to the active FeII state and retards the reaction. However, when the 1,10-phenanthroline is protonated, it acts as a hydrogen-bond donor which enhances O2 reduction by stabilizing FeIII-O2.- and FeIII-OOH intermediates and activating the O-O bond for cleavage. On the basis of these data, general guidelines for controlling the different possible rate-determining steps in the complex multistep 4e-/4H+ ORR are developed and a bioinspired principle-based design of an efficient electrochemical ORR is presented.

Bioinorganic Chemistry on Electrodes: Methods to Functional Modeling.

One of the major goals of bioinorganic chemistry has been to mimic the function of elegant metalloenzymes. Such functional modeling has been difficult to attain in solution, in particular, for reactions that require multiple protons and multiple electrons (nH+/ne-). Using a combination of heterogeneous electrochemistry, electrode and molecule design one may control both electron transfer (ET) and proton transfer (PT) of these nH+/ne- reactions. Such control can allow functional modeling of hydrogenases (H+ + e- → 1/2 H2), cytochrome c oxidase (O2 + 4 e- + 4 H+ → 2 H2O), monooxygenases (RR'CH2 + O2 + 2 e- + 2 H+ → RR'CHOH + H2O) and dioxygenases (S + O2 → SO2; S = organic substrate) in aqueous medium and at room temperatures. In addition, these heterogeneous constructs allow probing unnatural bioinspired reactions and estimation of the inner- and outer-sphere reorganization energy of small molecules and proteins.

Visualization of ceramide channels by transmission electron microscopy.

Functional studies have shown that the sphingolipid ceramide, self-assembles in phospholipid membranes to form large channels capable of allowing proteins to cross the membrane. Here these channels are visualized by negative stain transmission electron microscopy. The images contain features consistent with stain-filled pores having a roughly circular profile. There is no indication of tilt, and the results are consistent with the formation of right cylinders. The sizes of the pores range from 5 to 40nm in diameter with an asymmetric distribution indicating no apparent upper size limit. The size distribution matches well with the distribution of sizes calculated from electrophysiological measurements.

Role of pepsin in modifying the allergenicity of bhetki (Lates calcarifer) and mackerel (Rastrelliger kanagurta) fish.

The effect of pepsin digestion on the allergenicity of raw and thermally processed (boiled and fried) fish muscle extracts of two widely consumed fishes bhetki (Lates calcarifer) and mackerel (Rastrelliger kanagurta) was studied. Sere were collected from 110 patients who were hypersensitive to fish, as evidenced by their clinical history, symptoms and positive skin-prick test results. The various extracts after digestion with pepsin at different times of incubation were tested for specific IgE-binding activity by ELISA and immunoblotting with patients' sera. All the extracts of both the fishes retained their allergenicity as evidenced by ELISA and immunoblotting. In bhetki, maximum allergenicity was found in the pepsin-digested fried extract, whereas similar treatment decreased the allergenicity in fried mackerel. Results showed that raw as well as thermally processed allergens of both the fishes maintained strong allergenicity, even after digestion with pepsin for different time periods. The study revealed that the fish proteins played an important role in manifestation of allergy, due to their stable structure, which was retained even after pepsin and heat treatment.