A role for highly conserved carboxylate, aspartate-140, in oxygen activation and heme degradation by heme oxygenase-1.

Heme oxygenase (HO) catalyzes the oxygen-dependent degradation of heme to biliverdinIXalpha, CO, and free iron ion via three sequential monooxygenase reactions. Although the distinct active-site structure of HO from cytochrome P450 families suggests unique distal protein machinery to activate molecular oxygen, the mechanism and the key amino acid for the oxygen activation have not been clear. To investigate the functionality of highly conserved polar amino acids in the distal helix of HO-1, we have prepared alanine mutants: T135A, R136A, D140A, and S142A, and found drastic changes in the heme degradation reactions of D140A. In this paper, we report the first evidence that D140 is involved in the oxygen activation mechanism in HO-1. The heme complexes of HO mutants examined in this study fold and bind heme normally. The pK(a) values of the iron-bound water and autoxidation rates of the oxy-form are increased with R136A, D140A, and S142A mutations, but are not changed with T135A mutation. As the wild-type, T135A, R136A, and S142A degrade heme to verdohemeIXalpha with H(2)O(2) and to biliverdinIXalpha with the NADPH reductase system. On the other hand, D140A heme complex forms compound II with H(2)O(2), and no heme degradation occurs. For the NADPH reductase system, the oxy-form of D140A heme complex is accumulated in the reaction, and only 50% of heme is degraded. The stopped flow experiments suggest that D140A cannot activate iron-bound dioxygen and hydroperoxide properly. To investigate the carboxylate functionality of D140, we further replaced D140 with glutamic acid (D140E), phenylalanine (D140F), and asparagine (D140N). D140E degrades heme normally, but D140N shows reactivity similar to that of D140A. D140F loses heme degradation activity completely. All of these results indicate that the carboxylate at position 140 is essential to activate the iron-bound dioxygen and hydroperoxide. On the basis of the present findings, we propose an oxygen activation mechanism involving the hydrogen-bonding network through the bridging water and D140 side chain.     

Enantioselective addition of ketene silyl acetals to nitrones catalyzed by chiral titanium complexes. Synthesis of optically active beta-amino acids

A chiral titanium complex, Ti(O-i-Pr)(4)/BINOL/tert-butylcatechol, catalyzes enantioselective addition reaction of ketene silyl acetals to nitrones to give optically active beta-amino acid derivatives which are biologically active compounds and useful synthetic intermediates of natural products and pharmaceuticals such as beta-lactam antibiotics. The combined process of catalytic oxidation of secondary amines and enantioselective carbon-carbon bond formation of nitrones thus obtained with ketene silyl acetals provides a useful two-step method for the synthesis of optically active beta-amino acid derivatives and related nitrogen compounds.     

Quasi-particle structure around a single vortex in high-Tc superconductors

For analyzing the checker-board like modulation of the local density of states (LDOS) around a vortex observed in the slightly overdoped Bi₂Sr₂CaCu₂O_x, we examined the effect of pseudogap state of high-T_c superconductors to the LDOS around the vortex. We first derived the Bogoliubov-de Gennes equation for d-wave superconductivity (d-SC) in the presence of d-spin density wave (d-SDW). Using the Fourier–Bessel expansion, we solved this equation for a single vortex state, numerically. We found that the peak of the bound states around E = 0 becomes small and modulation of the LDOS is observed for larger d-SDW order parameter.     

Stroboscopic phase portrait and strange attractors

External periodic modulation of a nonlinear oscillator may lead to a chaotic output behaviour. This phenomenon is attributed to the existence of a strange attractor, which embodies essentially a folding motion as is met in Bernoulli shift or the Baker's transformation.     

Luminescence channels of manganese-doped spinel

Two independent luminescence channels are observed from manganese-doped spinel Mn:MgAl₂O₄. The luminescence around 520 nm is assigned to transition from the lowest electronic excited state ⁴T₁ to the ground state ⁶A₁ of Mn²⁺ (3d)⁵ ion by analyzing the excitation spectrum and electron spin resonance measurement. The emission at 650 nm is triggered by the band edge excitation and is assigned similarly to the charge-transfer process associated with the manganese ion.     

PNA for one-base differentiating protection of DNA from nuclease and its use for SNPs detection

By the combination of peptide nucleic acid (PNA) with single-stranded DNA specific nucleases, alteration of a single base to another in DNA has been detected with high accuracy. Only the DNAs in DNA/PNA duplexes involving a mismatch are efficiently hydrolyzed by these enzymes, whereas fully matching sequences are kept intact. This difference is visually scored by adding 3,3'-diethylthiadicarbocyanine, which changes its color from blue to purple upon binding to DNA/PNA duplexes. These findings are applied to the convenient and straightforward detection of single nucleotide polymorphisms (SNPs). When the target site in the sample DNA is completely complementary with the PNA, a notable amount of DNA/PNA duplex remains and thus the solution exhibits purple color. In the presence of even one mismatch between PNA and DNA, however, the DNA is completely digested by the enzyme and therefore the dye shows its intrinsic blue color. The SNPs in the apolipoprotein E gene of human DNA have been successfully genotyped by this method.     

Rayleigh instabilities in multiply charged sodium clusters

The stability of multiply charged sodium clusters Na(q+)(n) (q< or =10) produced in collisions between neutral clusters and multiply charged ions A(z+) ( z = 1 to 28) is experimentally investigated. Multiply charged clusters are formed within a large range of temperatures and fissilities. They are identified by means of a high-resolution reflectron-type time-of-flight mass spectrometer (m/deltam approximately 14 000). The maximum fissility of stable clusters is obtained for z = 28 and is X approximately 0.85+/-0.07, slightly below the Rayleigh limit (X = 1). It is mainly limited by the initial cluster temperature (T approximately 100 K).     

Measurement of interfluorine distances in solids

(19)F homonuclear dipolar recoupling methods were used to measure internuclear distances ranging from 5 to 12 A in fluorinated organic compounds in the solid state. Magic-angle-spinning-based high-resolution techniques were utilized. Trifluoromethyl and aromatic fluorine groups were separated by rigid aromatic spacers; these compounds were diluted into nonfluorinated host molecule matrices to give isolated homonuclear spin pairs with known internuclear distances. Radiofrequency-driven recoupling (RFDR) was used to elicit magnetization exchange between the spin pairs in 1D and 2D experiments. Simulation of the exchange was accomplished using a Monte Carlo-type algorithm to search the parameter space. These methods allow the determination of distances with an accuracy of 1 A at shorter distances and 2 A at longer distances, with the assumption of no prior knowledge of T(2)(ZQ).     

Characterization of the activity of L-ascorbic acid 2-[3,4-dihydro-2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridecyl)-2H-1-be nzopyran-6-yl-hydrogen phosphate] potassium salt in hydroxyl radical elimination

The effect of L-ascorbic acid 2-[3,4-dihydro-2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridecyl)-2H -1-benzopyran-6-yl-hydrogen phosphate] potassium salt (EPC-K1) on hydroxyl radical (*OH) elimination was studied using electron spin resonance (ESR) and spectrophotometric experiments. The addition of EPC-K, and *OH scavengers eliminated the *OH generated from Cu2+/H2O2, Fe2+/H2O2 and H2O2/UV-irradiation reaction systems. However, in competitive reactions using different concentrations of a spin-trap agent, the addition of the *OH scavenger altered the IC50 values, whereas the addition of EPC-K1 and a metal chelater did not change the value in the Cu2+/H2O2 and Fe2+/H2O2 reaction systems. The addition of EPC-K1 and metal chelater changed the ESR signal for free Cu2+. The spectrophotometric experiments confirmed that the addition of EPC-K1 and metal chelater altered the absorption spectra due to CuCl2 and FeSO4, whereas the *OH scavenger did not alter the spectra. Therefore, it was demonstrated that EPC-K, has the ability both to scavenge *OH directly and to inhibit the generation of *OH by the chelation of Cu2+ and Fe2+.