Hybrid microwave oscillators with a virtual cathode

A review is given of the developments and theoretical investigations of a fundamentally new class of microwave devices, namely, hybrid microwave oscillators with a virtual cathode, which combine the useful properties of virtual cathodes with the advantages of those traditional microwave oscillators that operate with subcritical-current beams and have a high efficiency in generating ultrarelativistic electron beams. Among such devices are the following: a hybrid diffractional microwave oscillator with a virtual cathode, a hybrid gyrodevice with a virtual cathode, a hybrid beam-plasma vircator, a hybrid gyrocon with a virtual cathode, a hybrid Cherenkov oscillator with a virtual cathode, a hybrid microwave oscillator of the “vircator + traveling-wave tube” type, an original two-beam tube with a virtual cathode, and a klystron-like vircator.     

Identification of the 7-hydroxymethyl chlorophyll a reductase of the chlorophyll cycle in Arabidopsis

The interconversion of chlorophyll a and chlorophyll b, referred to as the chlorophyll cycle, plays a crucial role in the processes of greening, acclimation to light intensity, and senescence. The chlorophyll cycle consists of three reactions: the conversions of chlorophyll a to chlorophyll b by chlorophyllide a oxygenase, chlorophyll b to 7-hydroxymethyl chlorophyll a by chlorophyll b reductase, and 7-hydroxymethyl chlorophyll a to chlorophyll a by 7-hydroxymethyl chlorophyll a reductase. We identified 7-hydroxymethyl chlorophyll a reductase, which is the last remaining unidentified enzyme of the chlorophyll cycle, from Arabidopsis thaliana by genetic and biochemical methods. Recombinant 7-hydroxymethyl chlorophyll a reductase converted 7-hydroxymethyl chlorophyll a to chlorophyll a using ferredoxin. Both sequence and biochemical analyses showed that 7-hydroxymethyl chlorophyll a reductase contains flavin adenine dinucleotide and an iron-sulfur center. In addition, a phylogenetic analysis elucidated the evolution of 7-hydroxymethyl chlorophyll a reductase from divinyl chlorophyllide vinyl reductase. A mutant lacking 7-hydroxymethyl chlorophyll a reductase was found to accumulate 7-hydroxymethyl chlorophyll a and pheophorbide a. Furthermore, this accumulation of pheophorbide a in the mutant was rescued by the inactivation of the chlorophyll b reductase gene. The downregulation of pheophorbide a oxygenase activity is discussed in relation to 7-hydroxymethyl chlorophyll a accumulation.     

Microbiological transformations of delta6a10a-tetrahydrocannabinol.

A screening program was conducted to find microorganisms that catalyze transformation reactions with cannabinoids. Three hundred fifty-eight cultures, consisting of 97 bacteria, 175 actinomycetes, and 86 molds, were incubated in media containing 0.5 mg of Delta(6a,10a)-tetrahydrocannabinol (Delta(6a,10a)-THC) per ml. After 120 h of cultivation, ethyl acetate extracts of the cultures were examined by thin-layer chromatography (TLC) for transformation products. About 18% of the cultures modified Delta(6a,10a)-THC. The ability to modify the substrate did not predominate among any particular group of microorganisms. After purification, the products from three cultures were analyzed by high-resolution mass spectrometry, 100-mHz proton magnetic resonance spectrometry, ultraviolet spectrometry, and infrared spectrometry. These spectral data indicated that a Mycobacterium sp. oxidized Delta(6a,10a)-THC to cannabinol and a diastereomeric pair of 6a-hydroxy-Delta(10,10a)-THC isomers; a Streptomyces sp. and a Bacillus sp. oxidized Delta(6a,10a)-THC to 7-keto-Delta(6a,10a)-THC and 4'-hydroxy-Delta(6a,10a)-THC, respectively. The occurrence of these products and the presence of others that have not yet been isolated or identified indicate that microbial transformation may be a useful tool for the preparation of new cannabinoids that have desirable pharmacological properties.     

Hierarchical graphs for rule-based modeling of biochemical systems

Background  

In rule-based modeling, graphs are used to represent molecules: a colored vertex represents a component of a molecule, a vertex attribute represents the internal state of a component, and an edge represents a bond between components. Components of a molecule share the same color. Furthermore, graph-rewriting rules are used to represent molecular interactions. A rule that specifies addition (removal) of an edge represents a class of association (dissociation) reactions, and a rule that specifies a change of a vertex attribute represents a class of reactions that affect the internal state of a molecular component. A set of rules comprises an executable model that can be used to determine, through various means, the system-level dynamics of molecular interactions in a biochemical system.     

Determination of Some Molecular Parameters of Tyrosine Hydroxylase From Rat Adrenal, Rat Striatum, and Human Pheochromocytoma

The molecular parameters of tyrosine hydroxylase (EC 1.14.16.2) from rat adrenal, rat striatum, and human pheochromocytoma were determined by combined gel filtration and sucrose gradient ultracentrifugation. The enzyme from rat adrenal has a calculated molecular weight of 228,000, a Stokes radius of 60.9 A, a sedimentation coefficient of 9.10S, and a frictional ratio of 1.39. The enzyme from rat striatum has a calculated molecular weight of 210,000, a Stokes radius of 54.3 A, a sedimentation coefficient of 9.38S, and a frictional ratio of 1.28. Tyrosine hydroxylase from human pheochromocytoma tissue has a calculated molecular weight of 255,000, a Stokes radius of 68.2 A, a sedimentation coefficient of 9.08S, and a frictional ratio of 1.50. These results indicate that the tyrosine hydroxylases from central and peripheral tissue in the rat are quite similar although the human enzyme appears to be significantly larger.     

Analysis of the binding of fluorescent C5a and C3a to human peripheral blood leukocytes

Fluorescein-labeled human C5a and C3a were prepared and utilized to analyze the binding of C5a and C3a to human neutrophils and mononuclear cells. The fluorescein derivatives of C5a (Fl-C5a) and C3a (Fl-C3a) contained approximately one fluorescein molecule per molecule of protein. Fl-C5a retained biologic activity as determined by neutrophil O2- production, enzyme release, receptor binding, and reaction with rabbit anti-C5a antibody. Fl-C3a was biologically active as measured by contraction of guinea pig ileal strips, and maintained 87% of its antigenic character when reacted with rabbit anti-human C3a. The binding of Fl-C5a and Fl-C3a to human neutrophils and mononuclear cells was assessed with the use of flow cytometry. Fl-C5a bound to greater than 90% of neutrophils, with an average ED50 ranging from 2.8 to 6.8 nM, depending on the method of analysis. Fl-C5a binding to neutrophils was specific and was not inhibited by the presence of formyl-methionyl-leucyl-phenylalanine (f-MLP), C3a, or casein. Fl-C5a binding was totally blocked by an excess of C5a. C5a des arg partially inhibited the binding of Fl-C5a to neutrophils, but was 1000-fold less effective than C5a. Similar experiments with mononuclear cells showed that Fl-C5a was bound by monocytes but not by lymphocytes. Fl-C5a binding to monocytes was blocked totally by C5a but not by C3a or f-MLP. Comparative binding studies with neutrophils, monocytes, and lymphocytes showed that Fl-C5a was bound by an average of 93% +/- 4 of neutrophils, 68% +/- 9 of monocytes, and 6% +/- 3 of lymphocytes. Fl-C3a did not show significant binding to neutrophils, monocytes, or lymphocytes. These studies demonstrate that fluorescein derivatives of C5a and C3a can be prepared with retention of biologic activity, and provide a means to evaluate the binding of C5a to individual cells.     

Amino acid sequences of a-factor mating peptides from Saccharomyces cerevisiae

The molecular structure of a-factor, the mating hormone produced by mating type a cells of Saccharomyces cerevisiae, has been investigated. In culture filtrates of a cells four oligopeptides (a1 to a4) exhibiting a-factor activity have been found. These peptides have been isolated and their amino acid sequences have been determined. The a-factor peptides comprise two (apparently identical) pairs, a1/a2 and a3/a4, which differ in an interchange at position 6 of a valine in a1/a2 for a leucine in a3/a4. a1 and a4, which can be obtained by oxidation with H2O2 of purified a2 and a3, respectively, obviously represent oxidation artifacts formed under the conditions of culture. The amino acid sequences determined for the a-factor peptides are Tyr-Ile-Ile-Lys-Gly-Val Leu-Phe-Trp-Asp-Pro-Ala-Cys. Several lines of evidence suggest that the carboxyl-terminal cysteine residue is S-alkylated by a hydrophobic moiety.     

Response of cytochrome a,a3 to carbon monoxide in canine hearts with prior infarcts

The effects of moderate levels of carbon monoxide (CO) on the oxidation-reduction state of cytochrome a,a3 (cyt a,a3) were examined in the hearts of twelve dogs with a prior myocardial infarction. Exposure to ten minutes CO produced a carboxyhemoglobin (CO-Hb) level of 9.4%, a level experienced by heavy smokers. Accompanying the exposure to CO, cyt a,a3 became more reduced; 17.4% +/- 4.7%. Exposure to CO was accompanied by an increase of 33% +/- 4% in the rate of cyt a,a3 reduction following occlusion of the left circumflex coronary artery and a decrease of 24% +/- 8% in the rate of cyt a,a3 oxidation with release. There was also a decrease in the magnitude of cyt a,a3 reduction from 86% +/- 9% to 70% +/- 11%. These results indicate that moderate levels of CO trap cyt a,a3 in the reduced state which impairs the ability of the heart to recover from transient ischemic episodes.     

Pathogenicity of two species of Fusarium on some cultivars of bean in greenhouse

Twenty isolates of Fusarium oxysporum and F. solani were isolated from the infected roots of bean in different farms of east Azarbaijan and Tehran Provinces and their pathogenicity determined. Most isolates of the fungi were identified as F. oxysporun. They caused root rot, yellowing and wilting of bean in the field. In this test, the roots of 6 cultivars of bean seedlings soaked in suspension of the 7 isolates of the fungi (a1, Gogan, a2, Bilverdi, a3, Savojbolagh-Hashtgerd, a4, field of Agr. Coll. a5, Khomein, a6, Ramjin of F. oxysporum and a7 of F. solani of Varamin, Iran) for 5 minute (106 spores/ml.) then transplanted into the sterilized soil in 4 pots (as replication). For control (a8) the roots soaked in distilled water. The results showed that percentage average of necrotic roots and crowns of isolates al, a2, a3, a5, a6, a7 was %20.31 in group a, a4 was %43.52 in group b and a8 was %2.77 in group c after 3 weeks. The isolate a4 (from the field of Agricultural College, Karaj) was more infectious than the other because it caused wilting, yellowing the leaves and decreased the growth very soon, followed by a5 with %25.32 rate was more pathogenic. Bean cultivar Goli-Red was more tolerant with %10.02 than the others of 16.29 (Naz Red) to 25.15 percent of necrotic the roots & stems.     

Fuzzy molecular surfaces.

Surface area of a macromolecule, accessible to a solvent, is defined and calculated, taking into account the probabilistic character of atomic positions due to the high frequency atomic vibrations. For a given a space point, we consider a probability of the event, that this point is covered by a macromolecule. A volume is defined as a space integral of this probability field. The envelope, accessible to a solvent molecule center, becomes fuzzy, existing only in a probabilistic sense. The accessible area is defined as a derivative of the envelope volume with respect to the probe size.     

The polymerase-like core of brome mosaic virus 2a protein, lacking a region interacting with viral 1a protein in vitro, maintains activity and 1a selectivity in RNA replication.

Brome mosaic virus (BMV), a member of the alphavirus-like super-family of positive-strand RNA viruses, encodes two proteins required for viral RNA replication: 1a and 2a. 1a contains m7G methyltransferase- and helicase-like domains, while 2a contains a polymerase (pol)-like core flanked by N- and C-terminal extensions. Genetic studies show that BMV RNA replication requires 1a-2a compatibility implying direct or indirect 1a-2a interaction in vivo. In vitro, la interacts with the N-terminal 125-amino-acid segment of 2a preceding the pol-like core, and prior deletion studies suggested that this 2a segment was essential for RNA replication. We have now used protein fusions and deletions to explore possible parallels between noncovalent 1a-2a interaction and covalent fusion of similar protein domains in tobacco mosaic virus and to see whether the N-terminal 2a-1a interaction was the primary basis for 1a-2a compatibility in vivo. We found that 2a can function as part of a tobacco mosaic virus-like 1a-2a fusion and that a 2a segment (amino acids 162 to 697) comprising the pol-like core was sufficient to provide 2a functions in such a fusion. Unexpectedly, the unfused 2a core segment also supported RNA replication when it and wild-type la were expressed as separate proteins. Moreover, in gene reassortant experiments with the related cowpea chlorotic mottle virus, the unfused 2a core segment showed the same 1a compatibility requirements as did wild-type BMV 2a. Thus, the pol-like core of 2a must interact with la in a way that is selective and essential for RNA synthesis, and 1a-2a interactions are more complex than the single, previously mapped interaction of the N-terminal 2a segment with 1a.     

Effects of Wnt proteins on cell proliferation and apoptosis in HEK293 cells

Wnt proteins and Wnt signalings have been implicated in a variety of development and cell processes, while aberrant activation of Wnt signaling is linked to a range of cancers in many tissues. In this study, we used the HEK293 cell line to investigate the effects of Wnt3a and Wnt5a on proliferation and apoptosis in a serum starvation culture. After Wnt3a and Wnt5a proteins were expressed, they both promoted the proliferation of HEK293 cells under serum starvation. After 48h of serum starvation, both Wnt3a and Wnt5a inhibited serum starvation-induced apoptosis of HEK293 cells and continued up to 96h. We demonstrated that Wnt3a and Wnt5a can promote proliferation of HEK293 cells and inhibit serum starvation-induced apoptosis, which implies that Wnt3a and Wnt5a can maintain the survival of HEK293 cells under stress, and also provide a novel insight into the role of Wnt3a and Wnt5a and their related signalings in carcinogenesis.     

ULTRASTRUCTURE AND SYSTEMATICS OF TWO NEW FRESHWATER RED CRYPTOMONADS, STOREATULA RHINOSA, SP. NOV. AND PYRENOMONAS OVALIS, SP. NOV.

The ultrastructure and systematics of two red colored freshwater cryptomonads, Storeatula rhinosa , sp. nov. and Pyrenomonas ovalis , sp. nov., are described for the first time. Storeatula, which had been described from marine waters only, has a single inner periplast sheet and a fibrous surface periplast component. Cells lack a furrow but possess a gullet, a bilobed chloroplast connected by a pyrenoid and a nucleomorph located in an indentation of the pyrenoid. This freshwater Storeatula possesses the same general features as the marine species, but it has a contractile vacuole and lacks the lobed chloroplast of S. major. P. ovalis has the generic characteristics described for marine species of Rhodomonas. These characteristics include a short furrow, a deep gullet, square inner periplast plates with beveled corners, a slightly fibrillar surface periplast component, a single chloroplast with two lobes connected by a pyrenoidal bridge and a nucleomorph located in an indentation of the pyrenoid.     

Molecular cloning and expression of the col2a1a and col2a1b genes in the medaka, Oryzias latipes

The Col2a1 gene is expressed in notochord, otic vesicle, cartilaginous tissue and the anlage of endochondral bone during development in higher vertebrates. Type II collagen, a homotrimeric product of the Col2a1 gene, functions as a key regulatory protein for cartilage development and endochondral ossification. In medaka and zebrafish, a single homolog of the col2a1 gene has been identified. However, it is necessary to note that many genes are duplicated in teleost fishes. To clarify function of col2a1 genes in teleost fishes and to further understand the process of cartilage development and endochondral ossification, we cloned and mapped the gene loci of two col2a1 orthologs in medaka. The proteins encoded by both medaka col2a1a and col2a1b genes were highly conserved (85.3% and 82.6%) relative to human COL2A1, but synteny was not observed. We also examined the expression patterns of col2a1a and col2a1b during embryonic development. Whole-mount insitu hybridization data suggests that expression patterns of both medaka co2a1a and col2a1b genes are similar to that of zebrafish co2a1 in the early embryonic stages. In medaka, the two col2a1 genes show a closely correlated pattern of spatial and temporal expression. In late embryonic stages, however, there were differences in both expression patterns in the pectoral fin. This study is the first report of two homologs of col2a1 in teleosts and also the first examination of col2a1a and col2a1b expression patterns in this group.     

Transient-state reduction and steady-state kinetic studies of menaquinol oxidase from Bacillus subtilis, cytochrome aa3-600 nm. Spectroscopic characterization of the steady-state species.

Cytochrome aa3-600 or menaquinol oxidase, from Bacillus subtilis, is a member of the heme-copper oxidase family. Cytochrome aa3-600 contains cytochrome a, cytochrome a3, and CuB, and each is coordinated via histidine residues to subunit I. Subunit II of cytochrome aa3-600 lacks CuA, which is a common feature of the cytochrome c oxidase family members. Anaerobic reduction of cytochrome aa3-600 by the substrate analogue 2,3-dimethyl-1,4-naphthoquinone (DMN) resolves two distinct kinetic phases by stopped-flow, single-wavelength spectrometry. Global analysis of time-resolved, multiwavelength spectra shows that during these distinct phases cytochromes a and a3 are both reduced. Cyanide binding to cytochrome a3 enhances the fast phase rate, which in the presence of cyanide can be assigned to cytochrome a reduction, whereas cytochrome a3-cyanide reduction is slow. The steady-state activity of cytochrome aa3-600 exhibits saturation kinetics as a function of DMN concentration with a Km of 300 microM and a maximal turnover of 63.5 s(-1). Global kinetic analysis of steady-state spectra reveals a species that is characteristic of a partially reduced oxygen adduct of cytochrome a3-CuB, whereas cytochrome a remains oxidized. Electron paramagnetic resonance (EPR) spectroscopy of the oxidase in the steady state shows the expected signal from ferricytochrome a, and a new EPR signal at g = 2.01. A model of the catalytic cycle for cytochrome aa3-600 proposes initial electron delivery from DMN to cytochrome a, followed by rapid heme to heme electron transfer, and suggests possible origins of the radical signal in the steady-state form of the enzyme.     

Multifaceted functions of RPS27a: An unconventional ribosomal protein

The ribosomal protein S27a (RPS27a) is cleaved from the fusion protein ubiquitin–RPS27a (Ub–RPS27a). Generally, Ub and RPS27a are coexpressed as a fusion protein but function independently after Ub is cleaved from RPS27a by a deubiquitinating enzyme. As an RP, RPS27a assembles into ribosomes, but it also functions independently of ribosomes. RPS27a is involved in the development and poor prognosis of various cancers, such as colorectal cancer, liver cancer, chronic myeloid leukemia, and renal carcinoma, and is associated with poor prognosis. Notably, the murine double minute 2/P53 axis is a major pathway through which RPS27a regulates cancer development. Moreover, RPS27a maintains sperm motility, regulates winged aphid indirect flight muscle degeneration, and facilitates plant growth. Additionally, RPS27a is a metalloprotein and mercury (Hg) biomarker. In the present review, we described the origin, structure, and biological functions of RPS27a.     

Characterization and expression pattern of zebrafish Anti-Müllerian hormone (Amh) relative to sox9a, sox9b, and cyp19a1a, during gonad development

The role of Anti-Müllerian hormone (Amh) during gonad development has been studied extensively in mammals, but is less well understood in other vertebrates. In male mammalian embryos, Sox9 activates expression of Amh, which initiates the regression of the Mullerian ducts and inhibits the expression of aromatase (Cyp19a1), the enzyme that converts androgens to estrogens. To better understand shared features of vertebrate gonadogenesis, we cloned amh cDNA from zebrafish, characterized its genomic structure, mapped it, analyzed conserved syntenies, studied its expression pattern in embryos, larvae, juveniles, and adults, and compared it to the expression patterns of sox9a, sox9b and cyp19a1a. We found that the onset of amh expression occurred while gonads were still undifferentiated and sox9a and cyp19a1a were already expressed. In differentiated gonads of juveniles, amh showed a sexually dimorphic expression pattern. In 31 days post-fertilization juveniles, testes expressed amh and sox9a, but not cyp19a1a, while ovaries expressed cyp19a1a and sox9b, but not amh. In adult testes, amh and sox9a were expressed in presumptive Sertoli cells. In adult ovaries, amh and cyp19a1a were expressed in granulosa cells surrounding the oocytes, and sox9b was expressed in a complementary fashion in the ooplasm of oocytes. The observed expression patterns of amh, sox9a, sox9b, and cyp19a1a in zebrafish correspond to the patterns expected if their regulatory interactions have been conserved with mammals. The finding that zebrafish sox9b and sox8 were not co-expressed with amh in oocytes excludes the possibility that amh expression in zebrafish granulosa cells is directly regulated by either of these two genes.     

Complete primary structure of human C4a anaphylatoxin

C4a anaphylatoxin is derived from the fourth component (C4) of the blood complement system. The C4 alpha-chain is selectively cleaved between positions 77 and 78 by the protease C1s, a subcomponent of C1, generating the fragments C4a and C4b. Human C4a was isolated directly from fresh serum after C1 of the classical pathway of complement was activated by heat-aggregated gamma-globulin. The C4a anaphylatoxin is a cationic polypeptide of Mr = 9000 composed of 77 residues and devoid of histidine, tryptophan, and carbohydrate. The primary structure of human C4a was deduced from sequence analysis of two cyanogen bromide fragments and of peptides obtained after chymotryptic digestion of the COOH-terminal cyanogen bromide fragment. The proposed sequence is: (formula, see text) Manual alignment of the linear structures of human C3a, C4a, and C5a, based primarily on the location of two Cys-Cys sequences in each indicate a 30% homology between C3a and C4a and a 36% homology between C5a and C4a. It was concluded from the sequence comparison that C3a, C4a, and C5a are a family of bioactive factors derived from precursor molecules that share a common genetic origin. Although the human anaphylatoxins share a partial structural identity and express similar biological activities, these factors ae immunologically distinct molecules having no antigenic determinants in common as judged by radioimmunoassay.