Synthesis of Samarium OxysulfateSm2O2SO4 in the High-Temperature Oxidation Reaction and Its Structural, Thermal and Luminescent Properties.

The oxidation process of samariumoxysulfide was studied in the temperature range of 500-1000 °C. Our DTA investigation allowed for establishing the main thermodynamic (∆Hºexp = -654.6 kJ/mol) and kinetic characteristics of the process (Ea = 244 kJ/mol, A = 2 × 1010). The enthalpy value of samarium oxysulfate (ΔHºf (Sm2O2SO4(monocl)) = -2294.0 kJ/mol) formation was calculated. The calculated process enthalpy value coincides with the value determined in the experiment. It was established that samarium oxysulfate crystallizes in the monoclinic symmetry class and its crystal structure belongs to space group C2/c with unit cell parameters a = 13.7442 (2), b = 4.20178 (4) and c = 8.16711 (8)Å, ß = 107.224 (1)°, V = 450.498 (9)Å3, Z = 4. The main elements of the crystalline structure are obtained and the cation coordination environment is analyzed in detail. Vibrational spectroscopy methods confirmed the structural model adequacy. The Sm2O2SO4luminescence spectra exhibit three main bands easily assignable to the transitions from 4G5/2 state to 6H5/2, 6H7/2, and 6H9/2 multiplets.

Relativistic transfer ionization and the Breit interaction.

We consider correlated transfer ionization in relativistic collisions between a highly charged ion and a light atom. In this process two quasifree electrons of the atom interact with each other during the short collision time that results in the capture of one of them by the ion and emission of the other. We show that this process is strongly influenced by the generalized Breit interaction already at modest relativistic impact energies.

Test of many-electron QED effects in the hyperfine splitting of heavy high-Z ions.

A rigorous evaluation of the two-photon exchange corrections to the hyperfine structure in lithiumlike heavy ions is presented. As a result, the theoretical accuracy of the specific difference between the hyperfine splitting values of H- and Li-like Bi ions is significantly improved. This opens a possibility for the stringent test of the many-electron QED effects on a few percent level in the strongest electromagnetic field presently available in experiments.

Structure of the wake of a magnetic obstacle.

We use a combination of numerical simulations and experiments to elucidate the structure of the flow of an electrically conducting fluid past a localized magnetic field, called magnetic obstacle. We demonstrate that the stationary flow pattern is considerably more complex than in the wake behind an ordinary body. The steady flow is shown to undergo two bifurcations (rather than one) and to involve up to six (rather than just two) vortices. We find that the first bifurcation leads to the formation of a pair of vortices within the region of magnetic field that we call inner magnetic vortices, whereas a second bifurcation gives rise to a pair of attached vortices that are linked to the inner vortices by connecting vortices.

[The dynamics of the direct current potentials of the brain during sustained attention in 10-11-year-old children].

The dynamics of direct current potentials of the brain was studied in 10-11-year-old children during sustained attention to successive presentation of series of Shulte tables. Children were examined twice: before and after the series of training to fast reading. A gradual increase in the level of direct current potentials during sustained attention was observed. The increase was more pronounced in children with excessive than in children with moderate reactions to the loading. After the series of training to fast reading, the increase in the level of direct current potentials was reduced in both groups. This aftertraining neurophysiological phenomenon was combined with a transformation of psychophysiological characteristics: a decrease in the time of viewing of Shulte tables and increase in the speed of reading. It is suggested that the shifts of direct current potentials reflects the dynamics of intensity of the cerebral energy metabolism.

The interdomain motions in myosin subfragment 1.

The interdomain motions in myosin subfragment 1 (S1) were studied by steady-state and time-resolved fluorescence of tryptophan residues and N-(iodoacetyl)-N'-(5-sulfo-1-naphtyl)ethylenediamine (AEDANS) attached to Cys178 of alkali light chain 1 (A1) exchanged into S1. The efficiency of fluorescence resonance energy transfer (FRET) from tryptophan residues of motor domain to AEDANS at A1 decreased dramatically after addition of ATP to S1A1-AEDANS. The efficiency of FRET calculated from the crystal structure of chicken S1 corresponded to the experimental one measured in the presence of ATP. The results showed that AEDANS at Cys178 of A1 became more mobile and distant from the motor domain of S1 upon ATP binding. These findings led to the suggestion that a release of the products of ATP hydrolysis and power stroke might be associated with movement of light chain-binding domain towards the N-terminal domain of S1.

[Two-dimensional reconstruction of birefringence map of the skeletal muscle sarcomere in relaxed and rigor states studied by interference microscopy]. / Dvumernaia rekonstruktsiia karty dvulucheprelomleniia sarcomera skeletnoi myshtsy v relaksirovannom i rigornom sostoianii po dannym interferentsionnoi mikroskopii.

A new method of automatized quantitative interferometry of skeletal muscle fibers was developed for the investigation of birefringence. A device based on the Linnic microscope was constructed to obtain phase images, which are two-dimensional pictures of birefringence. For the first time, two-and-three-dimensional maps of both total birefringence and birefringence for individual sarcomeres in the central part of muscle fiber were visualized using large databases. It was shown that total birefringence of fibers at rest length in the rigor state was lower as compared with the relaxed. Birefringence values from individual sarcomere interferograms revealed also that normalized A-disk birefringence was lower in the rigor state. The results obtained could be explained by a decrease of thick filaments anisotropy, due to the moving away of myosin heads from the rod during transition into the rigor state.

The identification of tryptophan residues responsible for ATP-induced increase in intrinsic fluorescence of myosin subfragment 1.

ATP binding to myosin subfragment 1 (S1) induces an increase in tryptophan fluorescence. Chymotryptic rabbit skeletal S1 has 5 tryptophan residues (Trp113, 131, 440, 510 and 595), and therefore the identification of tryptophan residues perturbed by ATP is quite complex. To solve this problem we resolved the complex fluorescence spectra into log-normal and decay-associated components, and carried out the structural analysis of the microenvironment of each tryptophan in S1. The decomposition of fluorescence spectra of S1 and S1-ATP complex revealed 3 components with maxima at ca. 318, 331 and 339-342 nm. The comparison of structural parameters of microenvironment of 5 tryptophan residues with the same parameters of single-tryptophan-containing proteins with well identified fluorescence properties applying statistical method of cluster analysis, enabled us to assign Trp595 to 318 nm, Trp440 to 331 nm, and Trp 13, 131 and 510 to 342 nm spectral components. ATP induced an almost equal increase in the intensities of the intermediate (331 nm) and long-wavelength (342 nm) components, and a small decrease in the short component (318 nm). The increase in the intermediate component fluorescence most likely results from an immobilization of some quenching groups (Met437, Met441 and/or Arg444) in the environment of Trp440. The increase in the intensity and a blue shift of the long component might be associated with conformational changes in the vicinity of Trp510. However, these conclusions can not be extended directly to the other types of myosins due to the diversity in the tryptophan content and their microenvironments.

[Analysis of log-normal components of fluorescence spectra of prodan and acrylodan bound to proteins]. / Analiz log-normal'nykh komponent spektrov fluorestsentsii sviazannykh s belkami prodana i akrilodana.

Steady-state fluorescence spectra of prodan and acrylodan covalently bound to cystein residue of Lys-Cys-Phe tripeptide in solvents of different polarity were analyzed. It was shown that the shape of spectral bands is well described by a log-normal function. Linear relations between three shape-determining parameters of the log-normal function (namely, the positions of spectral maximum and two half-maximum amplitudes) were revealed and evaluated for both fluorophores. This finding enabled us to present the shape of spectral bands of these fluorophores in any environment as analytical log-normal functions depending on only two parameters, the maximum position and the peak amplitude. The empirical uniparametric log-normal curve was used for the analysis of composite fluorescence spectra of prodan bound to bovine serum albumin and acrylodan covalently attached to actin or subfragment 1 of myosin.

Binding of myosin cross-bridges to thin filaments of rabbit skeletal muscle.

Cross-linking of myosin subfragment 1 (S1) with a molar excess of actin in vitro reveals the presence of an actin-S1-actin complex. It is absolutely essential that actin be present in molar excess over S1 so that the decoration of F-actin with S1 be incomplete. However, the excess of actin may not be available in the overlap zone of sarcomeres of skeletal muscle. We therefore found it necessary to test for the presence of the actin-S1-actin complex in vivo. Myofibrils from rabbit skeletal muscle were reacted with zero-length cross-linker, the products were resolved by polyacrylamide gel electrophoresis and analyzed by Western blots using antibodies against actin and against heavy and light chains of myosin. The cross-linking produced the evidence of formation of actin-S1-actin complex.

Interaction of the N-terminus of chicken skeletal essential light chain 1 with F-actin.

Skeletal myosin has two isoforms of the essential light chain (ELC), called LC1 and LC3, which differ only in their N-terminal amino acid sequence. The LC1 has 41 additional residues containing seven pairs of Ala-Pro, which form an elongated structure, and two pairs of lysines located near the N-terminus. When myosin subfragment-1 (S1) binds to actin, these lysines may interact with the C-terminus of actin and be responsible for the isoform specific properties of myosin. Here we employ cross-linking to identify the LC1 residues that are in contact with actin. S1 was reconstituted with various LC1 mutants and reacted with the zero-length cross-linker 1-ethyl-3-[3-dimethyl-aminopropyl]-carbodiimide (EDC). Cross-linking occurred only when actin was in molar excess over S1. Wild-type LC1 could be cross-linked through the terminal alpha-NH2 group, as well as via the two pairs of lysines. In a mutant ELC, where the lysines were deleted but two arginines were introduced near the N-terminus, the light chain could still be cross-linked via the terminal alpha-NH2 group. When the charge was reduced in the N-terminal region while retaining the Ala-Pro rich region, the mutant could not be cross-linked. These results suggest that as long as the N-terminus contains charged residues and an Ala-Pro rich extension, the binding between LC1 and actin can occur.

Insecticidal toxins from the bacterium Photorhabdus luminescens.

Transgenic plants expressing Bacillus thuringiensis (Bt) toxins are currently being deployed for insect control. In response to concerns about Bt resistance, we investigated a toxin secreted by a different bacterium Photorhabdus luminescens, which lives in the gut of entomophagous nematodes. In insects infected by the nematode, the bacteria are released into the insect hemocoel; the insect dies and the nematodes and bacteria replicate in the cadaver. The toxin consists of a series of four native complexes encoded by toxin complex loci tca, tcb, tcc, and tcd. Both tca and tcd encode complexes with high oral toxicity to Manduca sexta and therefore they represent potential alternatives to Bt for transgenic deployment.

Effect of ADP on binding of skeletal S1 to F-actin.

The proximity of skeletal myosin subfragment-1 (S1) to actin, and its orientation with respect to thin filaments of single muscle fibers, were compared in the presence and in the absence of ADP. The proximity was assessed by the efficiency of carbodiimide-induced cross-linking and the orientation by polarization of fluorescence of probes attached to the essential light chains. ADP made no difference in proximity or orientation when the molar ratio of S1 to actin was low or high. However, at the intermediate ratios, ADP made a significant difference. Strong dissociating agents, AMP-PNP and PPi, made significant differences at all ratios. To explain this behavior, it is unnecessary to invoke the ADP-induced "swinging" of the tail of S1. Rather, it is simply explained by the "two-state" model which we proposed earlier, in which S1 binds to one or to two actin protomers, depending on the saturation of the filaments with S1s. The dissociation induced by the ADP shifts the equilibrium between the two bound states. At high and low degrees of saturation, ADP is unable to significantly decrease the amount of S1 bound to F-actin. However, at intermediate saturation levels, ADP causes significantly more S1s to bind to two actins. These results suggest that the ADP-induced changes seen at the intermediate molar ratios are due to the dissociation-induced reorientation of S1.

Interaction of the heavy and light chains of cardiac myosin subfragment-1 with F-actin.

The interaction of the heavy chain (HC) and the light chain (cdLC1) of cardiac S1 (cdS1) with F-actin was studied by cross-linking, Western blotting, and fluorescence polarization methods. Incorporation of cdLC1 in cross-linked products was examined by Western blots using the primary antibody against 71-74 residues of cdLC1. Cross-linking with zero-length, water-soluble reagent yielded three products with apparent molecular masses of 150, 160, and 210 kD. Like in the case of cross-linking of skeletal S1 with actin, these complexes included only HC of S1 and actin. The composition of the products were as follows: 150 kD, one HC of S1 cross-linked through a primary site (on the C-terminal of the 20-kD fragment) to the N-terminus of actin; 160 kD, one HC of S1 cross-linked through a secondary site (on the 50 kD fragment) to the N-terminus of actin; and 210 kD, one HC of S1 cross-linked through primary and secondary sites to two actins. Four additional products with apparent molecular masses of 66, 120, 185, and 235 kD contained cdLC1 and were identified as cdLC1 + actin, cdLC1 + HCS1, cdLC1 + actin + HCS1, and cdLC1 + two actins + HCS1, respectively. The same products were observed when cross-linking was performed in cardiac myofibrils incubated with cdS1. The production of cross-linked complexes of the heavy and light chain with actin decreased with an increase in the molar ratio of cdS1:actin. To test whether the orientation of myosin heads depended on a degree of occupation of thin filaments, myofibrils were irrigated with varying concentrations of cdS1. Fluorescence polarization measurements of cdS1 bound to individual I-bands revealed that the orientation depended on the concentration.

Cytochrome P450 genes from Helicoverpa armigera: expression in a pyrethroid-susceptible and -resistant strain.

The molecular basis of metabolic resistance to pyrethroids in Helicoverpa armigera is currently under debate. Substantial indirect evidence supports a role for both esterase- and cytochrome-P450-mediated metabolism. However, the relative roles played by these two mechanisms in field-based resistance is uncertain. Our understanding of the importance of P450-mediated metabolism is hindered by the paucity of cloned genes from this species, and the corresponding absence of data on rates of insecticide metabolism by functionally expressed P450s. To facilitate P450 gene isolation from H. armigera we used degenerate primers in the reverse transcriptase-polymerase chain reaction (RT-PCR) to clone P450 gene fragments from the RNA of a pyrethroid-resistant strain. Here we report the isolation of eight new P450 genes: seven from the CYP4 family and one CYP9. One of these genes, CYP4G8, is two-fold over-expressed in the resistant strain, whereas the other CYP4s showed either similar or undetectable levels of expression. CYP9A3 appears to be a homolog of the putatively resistance-associated CYP9A1 of Heliothis virescens. However, no difference in expression between the H. armigera strains was detected. CYP6B2, a gene previously reported to be over-expressed in a different pyrethroid-resistant strain of H. armigera, also revealed non-detectable levels of expression in both strains. These observations suggest that different P450s may be over-expressed in different resistant strains, and emphasize that recombinant expression will be necessary in order to define precisely their individual substrate specificities and ability to metabolize pyrethroids. The gene fragments described here represent an important first step in this direction.

Molecular mapping of insecticide resistance genes in the yellow fever mosquito (Aedes aegypti).

Several loci conferring insecticide resistance in the yellow fever mosquito (Aedes aegypti) have previously been mapped by simple recombinational mapping. Here we describe correlation of these resistance phenotypes with molecular gene probes for insecticide target sites by RFLP mapping. The para sodium channel gene homologue and the GABA receptor gene Resistance to dieldrin map to the same genome regions as the DDT/pyrethroid and cyclodiene resistance loci, respectively. Although the acetylcholinesterase (target site of organophosphorus and carbamate insecticides) gene Ace does not map to any known resistance locus, it maps very close to the sex-determining locus. We discuss the possibilities that, if identified, Ace-mediated resistance in A. aegypti will be sex linked or that, as suggested for anopheline mosquitoes, two independent Ace loci may exist, one of which is autosomal. These results support the importance of target site insensitivity as an insecticide resistance mechanism in mosquitoes.

The sequence of a Drosophila Cyp4e2 cytochrome P450-encoding cDNA.

A composite 1458-bp cDNA that encodes cytochrome P450 (P450) Cyp4e2 has been constructed from clones isolated from two Drosophila embryonic cDNA libraries. The Drosophila cDNA open reading frame encodes a protein of 486 amino acids that is 40% identical and 61% similar to Cyp4d1 from Drosophila. The predicted protein is unusual in that it appears to lack the hydrophobic N-terminus typical of microsomal P450s and also contains a small insertion at its C-terminus.

Binding of S1(A1) and S1(A2) to F-actin.

The binding curve of myosin subfragment-1 (S1) to F-actin is not a simple hyperbola: at high concentrations of S1 the binding curve can be transformed into a linear plot ("normal" binding), but at small concentrations of S1 the binding complications deform the binding curve and produce nonlinear transforms ("anomalous" binding) [Andreev, O. A., & Borejdo, J. (1992) J. Muscle Res. Cell Motil. 13, 523-533]. This anomalous behavior may result either from the heterogeneity of S1 in regard to light chain isoforms or from the cooperativity between S1's. To distinguish between these possibilities we measured the affinity and the orientation of S1(A1) and S1(A2) with respect to F-actin. Affinity was measured in vitro by ultracentrifugation in the presence of F-actin, and orientation was measured in vivo by a combination of polarization of fluorescence and linear dichroism. We found that both the affinity and the orientation depended on the relative concentration of S1 isomer and actin: when S1 was in excess or was equimolar with actin (filament saturated with S1), each isomer bound F-actin with an affinity of 2 x 10(6) M-1 and was oriented approximately perpendicularly to the muscle axis. When actin was in excess (filament unsaturated with S1), each isomer bound F-actin with an affinity of 1.2 x 10(7) M-1 and was oriented more parallel to the muscle axis. S1(A1) and S1(A2) labeled on the light chain had different polarizations when bound to unsaturated filaments but had the same polarizations when bound to saturated filaments. These results excluded heterogeneity as a reason for anomalous binding and suggested that binding occurred with negative cooperativity. We think that the negative cooperativity occurs when saturation of actin filaments with heads leads to the lack of vacant adjacent sites on a filament and a consequent prevention of S1 binding to two actin protomers.

Binding of heavy-chain and essential light-chain 1 of S1 to actin depends on the degree of saturation of F-actin filaments with S1.

The interaction of heavy-chain isoforms of myosin subfragment-1 with actin was examined by cross-linking with carbodiimide (EDC). The heavy chain of S1 could be cross-linked to a single actin molecule through sites on either 20 or 50 kDa proteolytic domains, resulting in complexes which migrated in an 8% polyacrylamide gel in the presence of Tricine buffer with an apparent molecular mass (M(app)) of 150 or 160 kDa, respectively. Cross-linking of S1 through both sites to two actins produced a complex migrating with an M(app) of 210 kDa. Cross-linking of the S1(A1) isoform [but not S1(A2)] to F-actin produced four additional peptides with M(app) values of 64, 160, 185, 210, and 235 kDa complexes was almost inhibited at a high degree of saturation while the inhibition of the 150 kDa product was relatively small. At a low degree of saturation, the ratio of 150 to 160 kDa complexes was 1. Cross-linking between the S1 isoforms and regulated F-actin was not affected by Ca2+. These data show that contact of the S1 to one actin protomer is through a site on the 20 kDa fragment and to the second actin protomer through the sites located on the 50 kDa fragment and on the essential light-chain 1. At nonphysiological conditions of full saturation of actin filaments with myosin heads, the binding of heavy chain at S1 and of A1 to the second actin could be almost abolished.

Fluorescence polarization study of the rigor complexes formed at different degrees of saturation of actin filaments with myosin subfragment-1.

A serine residue located in the active site of myosin head (S1) was labelled by 9-anthroylnitrile, an amino group located in the central domain of S1 was labelled by 7-diethylamino-3-(4'-isothio-cyanato-phenyl)-4-methylcoumari n, a cysteine residue located near the C-terminus of S1 was labelled by 5-[2-((iodoacetyl)-amino)ethyl]-amino-naphthalene-1-sulfonic acid (1,5-IAEDANS) and a cysteine residue located near the C-terminus of the alkali light chain 1 was labelled with iodoacetamido-tetramethyl-rhodamine. Polarization of fluorescence of S1 was measured in solution (where it indicated the mobility of actin-bound S1) and in myofibrils (where it indicated orientation of probes) to check whether the anisotropy of S1 labelled at different positions depended on the molar ratio S1:actin. In solution, when increasing amounts of actin were added to a fixed amount of labelled S1 (i.e. when myosin heads were initially in excess over actin), anisotropy saturated at 1 mol of S1 per 1 mol of actin. When increasing amounts of S1 were added to a fixed amount of F-actin (i.e. when actin was initially in excess over S1), the anisotropy saturated at 1 mol of S1 per 2 mols of actin. In myofibrils, orientation of S1 was different when S1 was added at nanomolar concentration (intrinsic actin was in excess over extrinsic S1) then when it was added at micromolar concentration (excess of S1 over actin). The fact that the anisotropy of S1 labelled at different positions depended on the molar ratio excluded the possibility that changes were confined to one part of the cross-bridge and supports our earlier proposal that the two rigor complexes which S1 can form with F-actin differ globally in conformation.