Компьютерные технологии проведения практических занятий по электротехнике

The paper gives the rationale behind the methodological approach of conducting practical training on electrical engineering. The approach features extensive guidance on students ' preliminary extra-curricular work in the process of preparation for the classes (drawing up schemes and graphs, chain calculation, etc.) The extra-curricular work is followed by the computer analysis of the chain along with the comparison of the result

Influence of the Interdomain Interface on Structural and Redox Properties of Multiheme Proteins

Multiheme proteins are important in energy conversion and biogeochemical cycles of nitrogen and sulfur. A diheme cytochrome c4 (c4) was used as a model to elucidate roles of the interdomain interface on properties of iron centers in its hemes A and B. Isolated monoheme domains c4-A and c4-B, together with the full-length diheme c4 and its Met-to-His ligand variants, were characterized by a variety of spectroscopic and stability measurements. In both isolated domains, the heme iron is Met/His-ligated at pH 5.0, as in the full-length c4, but becomes His/His-ligated in c4-B at higher pH. Intradomain contacts in c4-A are minimally affected by the separation of c4-A and c4-B domains, and isolated c4-A is folded. In contrast, the isolated c4-B is partially unfolded, and the interface with c4-A guides folding of this domain. The c4-A and c4-B domains have the propensity to interact even without the polypeptide linker. Thermodynamic cycles have revealed properties of monomeric folded isolated domains, suggesting that ferrous (FeII), but not ferric (FeIII) c4-A and c4-B, is stabilized by the interface. This study illustrates the effects of the interface on tuning structural and redox properties of multiheme proteins and enriches our understanding of redox-dependent complexation

The Production of Nitrous Oxide by the Heme/Nonheme Diiron Center of Engineered Myoglobins (Fe_BMbs) Proceeds through a <i>trans</i>-Iron-Nitrosyl Dimer

Denitrifying NO reductases are transmembrane protein complexes that are evolutionarily related to heme/copper terminal oxidases. They utilize a heme/nonheme diiron center to reduce two NO molecules to N₂O. Engineering a nonheme Fe_B site within the heme distal pocket of sperm whale myoglobin has offered well-defined diiron clusters for the investigation of the mechanism of NO reduction in these unique active sites. In this study, we use FTIR spectroscopy to monitor the production of N₂O in solution and to show that the presence of a distal Fe_B^II is not sufficient to produce the expected product. However, the addition of a glutamate side chain peripheral to the diiron site allows for 50% of a productive single-turnover reaction. Unproductive reactions are characterized by resonance Raman spectroscopy as dinitrosyl complexes, where one NO molecule is bound to the heme iron to form a five-coordinate low-spin {FeNO}⁷ species with ν­(FeNO)_heme and ν­(NO)_heme at 522 and 1660 cm^–1, and a second NO molecule is bound to the nonheme Fe_B site with a ν­(NO)_FeB at 1755 cm^–1. Stopped-flow UV–vis absorption coupled with rapid-freeze-quench resonance Raman spectroscopy provide a detailed map of the reaction coordinates leading to the unproductive iron-nitrosyl dimer. Unexpectedly, NO binding to Fe_B is kinetically favored and occurs prior to the binding of a second NO to the heme iron, leading to a (six-coordinate low-spin heme-nitrosyl/Fe_B-nitrosyl) transient dinitrosyl complex with characteristic ν­(FeNO)_heme at 570 ± 2 cm^–1 and ν­(NO)_FeB at 1755 cm^–1. Without the addition of a peripheral glutamate, the dinitrosyl complex is converted to a dead-end product after the dissociation of the proximal histidine of the heme iron, but the added peripheral glutamate side chain in Fe_BMb2 lowers the rate of dissociation of the promixal histidine which in turn allows the (six-coordinate low-spin heme-nitrosyl/Fe_B-nitrosyl) transient dinitrosyl complex to decay with production of N₂O at a rate of 0.7 s^–1 at 4 °C. Taken together, our results support the proposed trans mechanism of NO reduction in NORs

Vibrational Analysis of Mononitrosyl Complexes in Hemerythrin and Flavodiiron Proteins: Relevance to Detoxifying NO Reductase

Flavodiiron proteins (FDPs) play important roles in the microbial nitrosative stress response in low-oxygen environments by reductively scavenging nitric oxide (NO). Recently, we showed that FMN-free diferrous FDP from <i>Thermotoga maritima</i> exposed to 1 equiv NO forms a stable diiron-mononitrosyl complex (deflavo-FDP­(NO)) that can react further with NO to form N₂O [Hayashi, T.; Caranto, J. D.; Wampler, D. A; Kurtz, D. M., Jr.; Moënne-Loccoz, P. Biochemistry 2010, 49, 7040−7049]. Here we report resonance Raman and low-temperature photolysis FTIR data that better define the structure of this diiron-mononitrosyl complex. We first validate this approach using the stable diiron-mononitrosyl complex of hemerythrin, Hr­(NO), for which we observe a ν­(NO) at 1658 cm^–1, the lowest ν­(NO) ever reported for a nonheme {FeNO}⁷ species. Both deflavo-FDP­(NO) and the mononitrosyl adduct of the flavinated FPD (FDP­(NO)) show ν­(NO) at 1681 cm^–1, which is also unusually low. These results indicate that, in Hr­(NO) and FDP­(NO), the coordinated NO is exceptionally electron rich, more closely approaching the Fe­(III)­(NO^–) resonance structure. In the case of Hr­(NO), this polarization may be promoted by steric enforcement of an unusually small FeNO angle, while in FDP­(NO), the Fe­(III)­(NO^–) structure may be due to a semibridging electrostatic interaction with the second Fe­(II) ion. In Hr­(NO), accessibility and steric constraints prevent further reaction of the diiron-mononitrosyl complex with NO, whereas in FDP­(NO) the increased nucleophilicity of the nitrosyl group may promote attack by a second NO to produce N₂O. This latter scenario is supported by theoretical modeling [Blomberg, L. M.; Blomberg, M. R.; Siegbahn, P. E. J. Biol. Inorg. Chem. 2007, 12, 79−89]. Published vibrational data on bioengineered models of denitrifying heme-nonheme NO reductases [Hayashi, T.; Miner, K. D.; Yeung, N.; Lin, Y.-W.; Lu, Y.; Moënne-Loccoz, P. Biochemistry 2011, 50, 5939−5947] support a similar mode of activation of a heme {FeNO}⁷ species by the nearby nonheme Fe­(II)

A Nonheme, High-Spin {FeNO}⁸ Complex that Spontaneously Generates N₂O

One-electron reduction of [Fe­(NO)-(N3PyS)]­BF₄ (<b>1</b>) leads to the production of the metastable nonheme {FeNO}⁸ complex, [Fe­(NO)­(N3PyS)] (<b>3</b>). Complex <b>3</b> is a rare example of a high-spin (<i>S</i> = 1) {FeNO}⁸ and is the first example, to our knowledge, of a mononuclear nonheme {FeNO}⁸ species that generates N₂O. A second, novel route to <b>3</b> involves addition of Piloty’s acid, an HNO donor, to an Fe^II precursor. This work provides possible new insights regarding the mechanism of nitric oxide reductases

The Hemophore HasA from <i>Yersinia pestis</i> (HasA_yp) Coordinates Hemin with a Single Residue, Tyr75, and with Minimal Conformational Change

Hemophores from <i>Serratia marcescens</i> (HasA_sm) and <i>Pseudomonas aeruginosa</i> (HasA_p) bind hemin between two loops, which harbor the axial ligands H32 and Y75. Hemin binding to the Y75 loop triggers closing of the H32 loop and enables binding of H32. Because <i>Yersinia pestis</i> HasA (HasA_yp) presents a Gln at position 32, we determined the structures of apo- and holo-HasA_yp. Surprisingly, the Q32 loop in apo-HasA_yp is already in the closed conformation, but no residue from the Q32 loop binds hemin in holo-HasA_yp. In agreement with the minimal reorganization between the apo- and holo-structures, the hemin on-rate is too fast to detect by conventional stopped-flow measurements

A Nonheme Iron(III) Superoxide Complex Leads to Sulfur Oxygenation

A new alkylthiolate-ligated nonheme iron complex, FeII(BNPAMe2S)Br (1), is reported. Reaction of 1 with O2 at −40 °C, or reaction of the ferric form with O2•– at −80 °C, gives a rare iron(III)-superoxide intermediate, [FeIII(O2)(BNPAMe2S)]+ (2), characterized by UV–vis, 57Fe Mössbauer, ATR-FTIR, EPR, and CSIMS. Metastable 2 then converts to an S-oxygenated FeII(sulfinate) product via a sequential O atom transfer mechanism involving an iron-sulfenate intermediate. These results provide evidence for the feasibility of proposed intermediates in thiol dioxygenases

Light-Induced N₂O Production from a Non-heme Iron–Nitrosyl Dimer

Two non-heme iron–nitrosyl species, [Fe₂(<i>N</i>‑Et‑HPTB)­(O₂CPh)­(NO)₂]­(BF₄)₂ (<b>1a</b>) and [Fe₂(<i>N</i>‑Et‑HPTB)­(DMF)₂(NO)­(OH)]­(BF₄)₃ (<b>2a</b>), are characterized by FTIR and resonance Raman spectroscopy. Binding of NO is reversible in both complexes, which are prone to NO photolysis under visible light illumination. Photoproduction of N₂O occurs in high yield for <b>1a</b> but not <b>2a</b>. Low-temperature FTIR photolysis experiments with <b>1a</b> in acetonitrile do not reveal any intermediate species, but in THF at room temperature, a new {FeNO}⁷ species quickly forms under illumination and exhibits a ν­(NO) vibration indicative of nitroxyl-like character. This metastable species reacts further under illumination to produce N₂O. A reaction mechanism is proposed, and implications for NO reduction in flavo­diiron proteins are discussed

Direct Observation of Oxygen Rebound with an Iron-Hydroxide Complex

The rebound mechanism for alkane hydroxylation was invoked over 40 years ago to help explain reactivity patterns in cytochrome P450, and subsequently has been used to provide insight into a range of biological and synthetic systems. Efforts to model the rebound reaction in a synthetic system have been unsuccessful, in part because of the challenge in preparing a suitable metal-hydroxide complex at the correct oxidation level. Herein we report the synthesis of such a complex. The reaction of this species with a series of substituted radicals allows for the direct interrogation of the rebound process, providing insight into this uniformly invoked, but previously unobserved process

Direct Observation of Oxygen Rebound with an Iron-Hydroxide Complex

The rebound mechanism for alkane hydroxylation was invoked over 40 years ago to help explain reactivity patterns in cytochrome P450, and subsequently has been used to provide insight into a range of biological and synthetic systems. Efforts to model the rebound reaction in a synthetic system have been unsuccessful, in part because of the challenge in preparing a suitable metal-hydroxide complex at the correct oxidation level. Herein we report the synthesis of such a complex. The reaction of this species with a series of substituted radicals allows for the direct interrogation of the rebound process, providing insight into this uniformly invoked, but previously unobserved process