Localization of the Enzymes Involved in the Photoevolution of H(2) from Acetate in Chlamydomonas reinhardtii

The localization of a series of enzymes involved in the anaerobic photodissimilation of acetate in Chlamydomonas reinhardtii F-60 adapted to a hydrogen metabolism was determined through the enzymic analyses of the chloroplastic, cytoplasmic, and mitochondrial fractions obtained with a cellular fractionation procedure that incorporated cell wall removal by treatment with autolysine, digestion of the plasmalemma with the detergent digitonin, and fractionation by differential centrifugation on a Percoll step gradient. The sequence of events leading to the photoevolution of H₂ from acetate includes the conversion of acetate into succinate via the extraplastidic glyoxylate cycle, the oxidation of succinate to fumarate by chloroplastic succinate dehydrogenase, and the oxidation of malate to oxaloacetate in the chloroplast by NAD dependent malate dehydrogenase. The level of potential activity for the enzymes assayed were sufficient to accommodate the observed rate of the photoanaerobic dissimilation of acetate and the photoevolution of H₂.     

Quantitation of the O(2)-Dependent, CO(2)-Reversible Component of the Postillumination CO(2) Exchange Transient in Tobacco and Maize Leaves

The postillumination transient of CO₂ exchange and its relation to photorespiration has been examined in leaf discs from tobacco (Nicotiana tabacum) and maize (Zea mays). Studies of the transients observed by infrared gas analysis at 1, 21, and 43% O₂ in an open system were extended using the nonsteady state model described previously (Peterson and Ferrandino 1984 Plant Physiol 76: 976-978). Cumulative CO₂ exchange equivalents (i.e. nanomoles CO₂) versus time were derived from the analyzer responses of individual transients. In tobacco (C₃), subtraction of the time course of cumulative CO₂ exchange under photorespiratory conditions (21 or 43% O₂) from that obtained under nonphotorespiratory conditions (1% O₂) revealed the presence of an O₂-dependent and CO₂-reversible component within the first 60 seconds following darkening. This component was absent in maize (C₄) and at low external O₂:CO₂ ratios (i.e. <100) in tobacco. The size of the component in tobacco increased with net photosynthesis as irradiance was increased and was positively associated with inhibition of net photosynthesis by O₂. This relatively simple and rapid method of analysis of the transient is introduced to eliminate some uncertainties associated with estimation of photorespiration based on the maximal rate of postillumination CO₂ evolution. This method also provides a useful and complementary tool for detecting variation in photorespiration.     

Some Enzymes and Properties of the Reductive Carboxylic Acid Cycle Are Present in the Green Alga Chlamydomonas reinhardtii F-60

The reductive carboxylic acid cycle, the autotrophic pathway of CO₂ assimilation in prokaryotes (photosynthetic and nonphotosynthetic autotrophic bacteria), was investigated in Chlamydomonas reinhardtii F-60, an algal mutant lacking a complete photosynthetic carbon reduction pathway (C₃) due to a deficiency in phosphoribulokinase. Evidence was obtained consistent with the presence of the reductive carboxylic acid cycle in F-60. This conclusion is based on the fact that: (a) acetate approximately doubled CO₂ fixation in whole cells (4 micromoles per milligram chlorophyll per hour) and in chloroplasts (32 nanomoles per milligram chlorophyll per hour); and (b) pyruvate synthase, α-ketoglutarate synthase, and ATP-citrate lyase, three indicators of the cycle, were found in cell-free extracts.     

Reinterpretation of the vibrational spectroscopy of the medicinal bioinorganic synthon c,c,t-[Pt(NH3)2Cl2(OH)2]

The Pt(IV) complex c,c,t-[Pt(NH₃)₂Cl₂(OH)₂] is an important intermediate in the synthesis of Pt(IV) anticancer prodrugs and has been investigated as an anticancer agent in its own right. An analysis of the vibrational spectroscopy of this molecule was previously reported (Faggiani et al., Can. J. Chem. 60:529, 1982), in which crystallographic determination of the structure of the complex permitted a site group approach. The space group, however, was incorrectly assigned. In the present study we have redetermined at high resolution crystal structures of c,c,t-[Pt(NH₃)₂Cl₂(OH)₂] and c,c,t-[Pt(NH₃)₂Cl₂(OH)₂]·H₂O₂, which makes possible discussion of the effect of hydrogen bonding on the N–H and O–H vibrational bands. The correct crystallographic site symmetry of the platinum complex in the c,c,t-[Pt(NH₃)₂Cl₂(OH)₂] structure is used to conduct a new vibrational analysis using both group-theoretical and modern density functional theory methods. This analysis reveals the nature and symmetry of the “missing band” described in the original publication and suggests a possible explanation for its disappearance.     

Production of extracellular H2O2 and L-lysine-α-oxidase during bulk growth of the fungus Trichoderma cf. aureoviride Rifai VKM F-4268D under salt stress

In this article, we report on Some physiological aspects of the synthesis of extracellular L-lysine-??-oxidase (LO) by the fungus Trichoderma cf. aureoviride Rifai VKM F-4268D under salt stress conditions and discuss the possible role of this enzyme for the producer. It has been shown that The synthesis of extracellular LO and proteolytic enzymes is induced in the fungus T. cf. aureoviride Rifai VKM F-4268D during submerged cultivation on wheat bran under salt stress. It has been shown that LO biosynthesis is accompanied by H₂O₂ accumulation in the growth medium. It seems that the extracellular LO synthesis followed by hydrogen peroxide production under stress conditions provides an adaptive advantage for the producer fungus in its competition with other organisms.     

[60]Fullerene displacement from fac-(dihapto-[60]fullerene)(dihapto-1,2-bis-(1,10-phenanthroline) tricarbonyl tungsten(0)

The Lewis bases triphenyl phosphine and tricyclohexyl phosphine (L) displace [60]fullerene (C₆₀) from fac-(η²-C₆₀)(η²-phen)W(CO)₃ (phen=1,10-phenanthroline) to produce fac-(η²-phen)(η¹-L)W(CO)₃. Under flooding conditions, the reactions were first order with respect to fac-(η²-C₆₀)(η²-phen)W(CO)₃. The order with respect to C₆₀ and L depends on the reaction conditions i.e., whether [C₆₀]/[L] ≈ 0 or 0?It [C₆₀]/[L] ≈ 1. Two limiting cases of an interchange displacement of [60]fullerene from fac-(η²-C₆₀)(η²-phen)W(CO)₃, whose relative contributions to the overall mechanism depend on the nature of the solvent, are proposed based on the rate law and on the activation parameters. The mechanism involves an initial [60]fullerene dissociation to produce (i) the electronically unsaturated intermediate (η²-phen)W(CO)₃ for the dissociative displacement and (ii) the solvated intermediate fac-(solvent)(η²-phen)W(CO)₃ for the solvent-assisted [60]fullerene dissociation. The W-C₆₀ bond energy in fac-(η²-C₆₀)(η²-phen)W(CO)₃ was estimated to be in the vicinity of 105 kJ/mol based on the enthalpy of activation of the step where presumably [60]fullerene dissociates from fac-(η²-C₆₀)(η²-phen)W(CO)₃ to produce (η²-phen)W(CO)₃.     

Synthesis and in vivo evaluation of [18F]2-(4-(4-(2-(2-fluoroethoxy)phenyl)piperazin-1-yl)butyl)-4-methyl-1,2,4-triazine-3,5(2H,4H)-dione ([18F]FECUMI-101) as an imaging probe for 5-HT1A receptor agonist in nonhuman primates

The 5-HT_1AR partial agonist PET radiotracer, [¹¹C]CUMI-101, has advantages over an antagonist radiotracer as it binds preferentially to the high affinity state of the receptor and thereby provides more functionally meaningful information. The major drawback of C-11 tracers is the lack of cyclotron facility in many health care centers thereby limiting widespread clinical or research use. We identified the fluoroethyl derivative, 2-(4-(4-(2-(2-fluoroethoxy)phenyl)piperazin-1-yl)butyl)-4-methyl-1,2,4-triazine-3,5(2H,4H)dione (FECUMI-101) (K_i = 0.1 nM; E_max = 77%; EC₅₀ = 0.65 nM) as a partial agonist 5-HT_1AR ligand of the parent ligand CUMI-101. FECUMI-101 is radiolabeled with F-18 by O-fluoroethylation of the corresponding desmethyl analogue (1) with [¹⁸F]fluoroethyltosylate in DMSO in the presence of 1.6 equiv of K₂CO₃ in 45 ± 5% yield (EOS). PET shows [¹⁸F]FECUMI-101 binds specifically to 5-HT_1AR enriched brain regions of baboon. The specificity of [¹⁸F]FECUMI-101 binding to 5-HT_1AR was confirmed by challenge studies with the known 5-HT_1AR ligand WAY100635. These findings indicate that [¹⁸F]FECUMI-101 can be a viable agonist ligand for the in vivo quantification of high affinity 5-HT_1AR with PET.     

Nitrogen Fixation (C(2)H(2) Reduction) by Broad Bean (Vicia faba L.) Nodules and Bacteroids under Water-Restricted Conditions

Water potentials of leaves and nodules of broad bean (Vicia faba L.) cultivated on a sandy mixture were linearly and highly (r² = 0.99) correlated throughout a water deprivation of plants. A decrease of 0.2 megapascal of the nodule water potential (Ψ_nod) induced an immediate 25% inhibition of the highest level of acetylene reduction of broad bean nodules attached to roots. This activity continued to be depressed when water stress increased, but the effect was less pronounced. Partial recovery of optimal C₂H₂ reduction capacity of mildly water stressed nodules (Ψ_nod = −1.2 megapascals) was possible by increasing the external O₂ partial pressure up to 60 kilopascals. The dense packing of the cortical cells of nodules may be responsible for the limitation of O₂ diffusion to the central tissue. Bacteroids isolated from broad bean nodules exhibited higher N₂ fixation activity with glucose than with succinate as an energy-yielding substrate. Bacteroids from stressed nodules appeared more sensitive to O₂, and their optimal activity declined with increasing nodule water deprivation. This effect could be partly due to decreased bacteroid respiration capacity with water stress. Water stress was also responsible for a decrease of the cytosolic protein content of the nodule and more specifically of leghemoglobin. The alteration of the bacteroid environment appears to contribute to the decline in N₂ fixation under water restricted conditions.     

Syntheses of 6-(2-hydroxy-6-phenylhexyl)-5,6-dihydro-2H-pyran-2-one derivatives and their cytotoxicities

The cytotoxicities against cancer cells (HL-60, HeLa) and insect cells (Sf9) of four stereoisomers of 6-(2-hydroxy-6-phenylhexyl)− 5,6-dihydro-2H-pyran-2-one (1) were evaluated, and then their structure-activity relationships examined. The 2′-dehydroxy derivative 5 of (6 R,2′R)- and (6 R,2′S)-1 showed the highest activity against HeLa cells (IC₅₀ = 1.4 μM). To evaluate the effect of the 2′-hydroxy group of 1, 6R-and 6S-oxetane derivatives were also synthesized and their activities examined. Against HeLa and HL-60 cells, the activities of the less potent stereoisomers were enhanced 3–4-fold by the introduction of the oxetane moieties at the 2′-position. Against the insect cell line (Sf9), phenyl derivative 7 showed the highest activity with an IC₅₀ value of 8.0 μM.     

Growth of a Strictly Anaerobic Bacterium on Furfural (2-Furaldehyde)

A strictly anaerobic bacterium was isolated from a continuous fermentor culture which converted the organic constituents of sulfite evaporator condensate to methane and carbon dioxide. Furfural is one of the major components of this condensate. This furfural isolate could degrade furfural as the sole source of carbon and energy in a defined mineral-vitamin-sulfate medium. Acetic acid was the major fermentation product. This organism could also use ethanol, lactate, pyruvate, or fumarate and contained cytochrome c₃ and desulfoviridin. Except for furfural degradation, the characteristics of the furfural isolate were remarkably similar to those of the sulfate reducer Desulfovibrio gigas. The furfural isolate has been tentatively identified as Desulfovibrio sp. strain F-1.     

Biosynthesis of the Fungal Estrogen F-2 and a Naturally Occuring Derivative (F-3) by Fusarium moniliforme

The fungal metabolites, F-2 and F-3, associated with estrogenism in swine, are produced by some races of Fusarium moniliforme isolated from toxic feeds.     

Adaptation of Chlamydomonas reinhardtii High-CO(2)-Requiring Mutants to Limiting CO(2)

Photosynthetic characteristics of four high-CO₂-requiring mutants of Chlamydomonas reinhardtii were compared to those of wild type before and after a 24-hour exposure to limiting CO₂ concentrations. The four mutants represent two loci involved in the CO₂-concentrating system of this unicellular alga. All mutants had a lower photosynthetic affinity for inorganic carbon than did the wild type when grown at an elevated CO₂ concentration, indicating that the genetic lesion in each is expressed even at elevated CO₂ concentrations. Wild type and all four mutants exhibited adaptive responses to limiting CO₂ characteristic of the induction of the CO₂-concentrating system, resulting in an increased affinity for inorganic carbon only in wild type. Although other components of the CO₂-concentrating system were induced in these mutants, the defective component in each was sufficient to prevent any increase in the affinity for inorganic carbon. It was concluded that the genes corresponding to the ca-1 and pmp-1 loci exhibit at least partially constitutive expression and that all components of the CO₂-concentrating system may be required to significantly affect the photosynthetic affinity for inorganic carbon.     

Optical activity of dimolybdenum complex induced by chiral,chelating diamine ligand; syntheses and structures of [Mo2(O2CCF3)2(S,S-dach)2(CH3CN)2][BF4]2 and [Mo2(O2CCF3)2(R,R-dach)2(CH3CN)2][BF4]2 (dach=1,2-diaminocyclohexane)

The first structurally characterized, quadruply bonded complexes containing chiral diamine ligands, [Mo₂(O₂CCF₃)₂(S,S-dach)₂(CH₃CN)₂][BF₄]₂ (1), and [Mo₂(O₂CCF₃)₂(R,R-dach)₂(CH₃CN)₂][BF₄]₂ (2); (dach=1,2-diaminocyclohexane) were prepared by reactions of [Mo₂(O₂CCF₃)₂(CH₃CN)₆][BF₄]₂ with S,S-dach and R,R-dach, respectively, in CH₃CN. Their UV–Vis and circular dichroism (CD) spectra have been recorded and their structures determined by X-ray crystallography. Crystals of complexes 1 and 2 conform to the space groups P2 with two independent half molecules in the asymmetric unit. The two molecules have a similar structure consisting of a Mo₂ unit bridged by two cis-trifluoroacetate ligands and chelated by two dach ligands. Two acetonitrile molecules are coordinated to the Mo centers along the MoMo bond. The absorption wavelength at 507 nm for both 1 and 2 can be assigned to δ_xy→δ_xy* transitions. The solution CD spectra of these two complexes show two prominent bands at 525 and 385 nm and form mirror images of each other. The solid CD spectra of complexes 1 and 2 show marked red-shift in the absorption energies as compared with those measured in solution. The one-electron static coupling mechanism was invoked to explain the CD spectra for these complexes and the second lowest energy bands were assigned to be δ_xy→δ_x²−y² transitions.     

Crystal structure of bis(pyrrole-N-carbothioate)bis(triphenylphosphine)nickel(II), C46H38N2S2P2Ni

The crystal structure of Ni(C₄H₄NCOS)₂[(C₆H₅)₃P]₂, Ni(ptc)₂(Ph₃P)₂, has been determined by single crystal X-ray diffraction methods. This species exists as a square planar complex. The structure is monoclinic, P2₁/n, a=12.759(4), b=10.069(2), c= 16.158(6) Å, and β=91.96(3)°. The unit cell contains 2 formula units with an observed density of 1.31 g cm⁻³ (1.34 calculated). The final R index= 0.047 (R_w=0.038) for 2618 non-zero reflections having I > 3 σ(I).     

E2F-1 binding affinity for pRb is not the only determinant of the E2F-1 activity

E2F-1 is the major cellular target of pRB and is regulated by pRB during cell proliferation. Interaction between pRB and E2F-1 is dependent on the phosphorylation status of pRB. Despite the fact that E2F-1 and pRB have antagonistic activities when they are overexpressed, the role of the E2F-1-pRB interaction in cell growth largely remains unknown. Ideally, it would be better to study the properties of a pRB mutant that fails to bind to E2F, but retains all other activities. To date, no pRB mutation has been characterized in sufficient detail to show that it specifically eliminates E2F binding but leaves other interactions intact. An alternative approach to this issue is to ask whether mutations that change E2F proteins binding affinity to pRB are sufficient to change cell growth in aspect of cell cycle and tumor formation. Therefore, we used the E2F-1 mutants including E2F-1/S332-7A, E2F-1/S375A, E2F-1/S403A, E2F-1/Y411A and E2F-1/L132Q that have different binding affinities for pRB to better understand the roles of the E2F-1 phosphorylation and E2F-1-pRB interaction in the cell cycle, as well as in transformation and gene expression. Data presented in this study suggests that in vivo phosphorylation at amino acids 332-337, 375 and 403 is important for the E2F-1 and pRB interaction in vivo. However, although E2F-1 mutants 332-7, 375 and 403 showed similar binding affinity to pRB, they showed different characteristics in transformation efficiency, G₀ accumulation, and target gene experiments.     

Homogeneous CO hydrogenation and transfer hydrogenation catalyzed by ruthenium(II) complexes containing optically active Ph2PCH(Ph)CH(Me)NH2 and 1,2-C5H8(PPh2)2 ligands

Combination of (1S,2S)-cyclopentanediylbis(diphenylphosphine) with [Ru(η⁴-C₈H₁₂){η³-(CH₂)₂CMe}₂] afforded the chelate complex [Ru{η³-(CH₂)₂CMe}₂{(1S,2S)-C₅H₈(PPh₂)₂}] (1), which gave (OC-6-13)-[RuCl₂{(1S,2S)-C₅H₈(PPh₂)₂}{(1S,2S)-Ph₂PCH(Ph)CH(Me)NH₂}] (2) upon reaction with methanolic HCl in acetone, followed by the addition of the β-aminophosphine in DMF. The (P ∩ N)₂-chelated complexes (OC-6-13)-[RuCl₂{(1S,2S)-Ph₂PCH(Ph)CH(Me)NH₂}₂] (3) and (OC-6-13)-[RuCl₂{(1R,2S)-Ph₂PCH(Ph)CH(Me)NH₂}₂] (4) resulted from RuCl₃ · 3H₂O and the P,N ligands under reducing conditions. The crystal structures of 3 and 4 were determined by single-crystal X-ray diffraction. Following activation by KOBu-t in isopropanol, compounds 2–4 catalyzed the enantioselective transfer hydrogenation of acetophenone with i-PrOH as the hydrogen source as well as the direct hydrogenation of the ketone by H₂ in low to moderate e.e. (up to 67%).     

Conformational effect of a spin crossover iron(II) complex: Bis[N-(2-methylimidazol-4-yl)methylidene-2-aminoethyl]propanediamineiron(II) perchlorate

A iron(II) complex of the linear hexadentate N₆ ligand H₂L^2-3-2, [Fe(H₂L^2-3-2)](ClO₄)₂, was synthesized and the spin crossover properties were investigated, where H₂L^2-3-2 denotes bis[N-(2-methylimidazol-4-yl)methylidene-2-aminoethyl]propanediamine. The complex showed a gradual and reversible one-step spin crossover (SCO) between the high-spin (S = 2) and low-spin (S = 0) states at T_1/2 = 208 K without hysteresis. The crystal structures were determined at 296 and 250 K (HS state), 230, 210, and 200 K (intermediate between the HS and LS states) and 150 and 110 K (LS state). The spin transition from 296 to 150 K accompanies with the conformation change of the chelate rings at the triamine moiety and the formation of the hydrogen bond network in the same space group of orthorhombic Pbcn (no. 60). However, in the LS state at 110 K, the space group changed from orthorhombic Pbcn at 150 K (Pcan when the same axial setting to 110 K was used) to monoclinic P2₁/a (no. 14) at 110 K, although no spin transition and no change of assembly structure between 150 and 110 K were observed. It give us an idea that the space group transformation is mainly related to the conformational thermodynamic stability of the chelate rings at the triamine moiety and is not directly correlated with the spin transition.