Analogues of NADP+ as inhibitors and coenzymes for NADP+ malic enzyme from maize leaves

Structural analogues of the NADP⁺ were studied as potential coenzymes and inhibitors for NADP⁺ dependent malic enzyme from Zea mays L. leaves. Results showed that 1, N⁶-etheno-nicotinamide adenine dinucleotide phosphate ( NADP⁺), 3-acetylpyridine-adenine dinucleotide phosphate (APADP⁺), nicotinamide-hypoxanthine dinucleotide phosphate (NHDP⁺) and -nicotinamide adenine dinucleotide 2: 3-cyclic monophosphate (23NADPc⁺) act as alternate coenzymes for the enzyme and that there is little variation in the values of the Michaelis constants and only a threefold variation in V_max for the five nucleotides. On the other hand, thionicotinamide-adenine dinucleotide phosphate (SNADP⁺), 3-aminopyridine-adenine dinucleotide phosphate (AADP⁺), adenosine 2-monophosphate (2AMP) and adenosine 2: 3-cyclic monophosphate (23AMPc) were competitive inhibitors with respect to NADP⁺, while -nicotinamide adenine dinucleotide 3-phosphate (3NADP⁺), NAD⁺, adenosine 3-monophosphate (3AMP), adenosine 2: 5-cyclic monophosphate (25AMPc), 5AMP, 5ADP, 5ATP and adenosine act as non-competitive inhibitors. These results, together with results of semiempirical self-consistent field-molecular orbitals calculations, suggest that the 2-phosphate group is crucial for the nucleotide binding to the enzyme, whereas the charge density on the C₄ atom of the pyridine ring is the major factor that governs the coenzyme activity.Abbreviations NADP⁺ 1, N⁶-etheno-nicotinamide adenine dinucleotide phosphate - NHDP⁺ nicotinamide-hypoxanthine dinucleotide phosphate - APADP⁺ 3-acetylpyridine-adenine dinucleotide phosphate - SNADP⁺ thionicotinamide-adenine dinucleotide phosphate - AADP⁺ 3-aminopyridine-adenine dinucleotide phosphate - 23NADPc⁺ -nicotinamide adenine dinucleotide 2: 3-cyclic monophosphate - 3NADP⁺ -nicotinamide adenine dinucleotide 3-phosphate - 2AMP adenosine 2-monophosphate - 3AMP adenosine 3-monophosphate - 23AMPc adenosine 2: 3 monophosphate cyclic - A adenosine - RuBP ribulose 1,5-bisphosphate - SCF-MO Self-Consistent Field-Molecular Orbitals (method)     

Synthesis of oligoguanylates on oligocytidylate templates

Summary Short oligocytidylates can act as templates for the self-condensation of guanosine 5-phosphorimidazolide. In the absence of a catalytic metal ion or in the presence of Pb²⁺ a noticeable template effect is already observed with the dimer and the yield of long oligomers reaches a plateau with a hexamer template. Short templates give oligomers longers than the template length. The products are predominantly 2-5 linked for the Pb²⁺-catalyzed reaction while mixed linkages are observed in the uncatalyzed reaction.In the presence of Zn²⁺, a template effect is first observed with the pentamer and is maximal by the heptamer. The products are predominantly 3-5 linked. Oligomers shorter than or as long as the template are obtained in substantial yield, and longer products in much lower yields.Abbreviations G Guanosine - Gp guanosine 2(3)-phosphate - pG guanosine 5-phosphate - Gp! guanosine cyclic 2,3-phosphate - ImpG guanosine 5-phosphorimidazolide - ImpG^* [8-¹⁴C]-guanosine 5-phosphorimidazolide - pGp 5-phosphoguanosine 2(3)-phosphate - G²pG guanylyl-[2-5]-guanosine - G³pG guanylyl-[3-5]-guanosine - ImpGpG 5-phosphorimidazolide of GpG - (pG)_n (n = 2,3) oligomers of pG - GppG P₁, P₂-diguanosine 5-diphosphate - GppGpG 5-[guanosine 5-pyrophosphate] of GpG - NH₂pG guanosine 5-phosphoramidate - (pG)₄₊ tetramer and higher oligoguanylates with 5 terminal phosphate - oligo(G) oligoguanylate - Cp cytidine 2(3)-phosphate - Cp! cytidine cyclic 2,3-phosphate - (Cp)_n–1 Cp! (n= 2,3,4) oligocytidylates terminated by 5-OH groups and 2,3-cyclic phosphates - oligo(C) oligocytidylate - poly(C) polycytidylic acid - poly(U) polyuridylic acid - poly(C,G) random copolymer of C and G - BAP bacterial alkaline phosphatase (E. coli) - EDTA ethylenediaminetetraacetic acid - R_f chromatographic mobility     

Poly(U)-directed peptide-bond formation from the 2′(3′)-glycyl esters of adenosine derivatives

Summary The self-condensation of 2(3)-O-glycyl esters of adenosine, adenosine-5-(O-methylphosphate) and P₁, P₂-diadenosine-5-pyrophosphate in 6.2 mM solutions at pH 8.0 and -5°C in the presence of 12.5 mM poly(U) yields approximately 3 times as much diketopiperazine as reactions without poly(U). As the concentration of 2(3)-O-(glycyl)-P₁, P₂-diadenosine-5-pyrophosphate is decreased from 6.2 mM to 1.5 mM the yield of diketopiperazine in the presence of poly(U) decreases slightly from 6.6% to 5.2%, whereas, in the absence of poly(U) the yield of diketopiperazine decreases substantially from 2.4% to 0.75%. The enhanced yield of diketopiperazine that is attributed to the template action of poly(U) is temperature dependent and is observed only at temperatures below 10°C (5°C to -5°C) for 6.2 mM 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate) and below 23°C (15°C to -5°C) for 6.2 mM 2(3)-O-(glycyl)-P₁, P₂-diadenosine-5-pyrophosphate. The absence of a template effect at high temperatures is attributed to the melting of the organized helices. The hydrolysis half-lives at pH 8.0 and -5°C of 2(3)-O-(glycyl)-adenosine, 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate), 2(3)-O-(glycyl)-P₁, P₂-diadenosine-5-pyrophosphate, and 5-O-(glycyl)-adenosine in the presence of poly(U) are substantially larger than their half-lives in the absence of poly(U). The condensation of 2(3)-O-(glycyl)-adenosine yields 5% of 5-O-(glycyl)-adenosine in the presence of poly(U) compared to 0.7% in the absence of poly(U).Abbreviations DKP diketopiperazine - (gly)₂ glycylglycine - (gly)₃ glycylglycylglycine - AppA-gly 2(3)-O-(glycyl)-P₁, P₂-diadenosine-5-pyrophosphate - MepA-gly 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate) - Ado-2(3)-gly 2(3)-O-(glycyl)-adenosine - Ado-5-gly 5-O-(glycyl)-adenosine - Boc-gly N-tert-butyloxycarbonylglycine - AppA P₁, P₂-diadenosine-5-pyrophosphate - MepA adenosine-5-(O-methylphosphate) - AppA-Boc-gly 2(3)-O-(Boc-glycyl)-P₁, P₂-diadenosine-5-pyrophosphate - Ado-5-Boc-gly 5-O-(Boc-glycyl)-adenosine - Ado-2(3)-Boc-gly 2(3)-O-(Boc-glycyl)-adenosine     

The influence of increasing chlorine content on the accumulation and metabolism of polychlorinated biphenyls (PCBs) by Paul's Scarlet Rose cells

Four radiolabled congeners of biphenyls with increasing chlorine content (biphenyl; 1-monochlorobiphenyl; 2,2,4,4-tetrachlorobiphenyl; and 2,2,4,4,5,5-hexachlorobiphenyl) were provided to suspension cultures of rose (Rosa sp. cv. Paul's Scarlet) for 4 days. Both the kinetics of ¹⁴C exchange between the cells and medium, and the metabolism of the parent compounds depended on the chlorine content of the congeners. Analysis of both the cells and their medium showed that of the recovered radioactivity 88%, 86%, and 3% of the biphenyl, 1-PCB, and 2,2,4,4-PCB were metabolized respectively to polar and insoluble residue products. The 2,2,4,4,5,5-PCB did not appear to be metabolized.     

Effects of midazolam on the contraction and relaxation of segments of thoracic aorta stripped of endothelium and stimulated by adrenaline – experimental study in rabbits

To evaluate the effects of midazolam on the angiokinesis of segments of rabbits' thoracic aorta stripped of endothelium and stimulated by adrenaline.Two groups of aortic rings removed from albinic rabbits anesthetized with thiopental were used (Group I – 6 animals; Group II – 12 animals), stripped of endothelium, studied in an organ chamber, perfused by Krebs-Henseleit solution. The groups were stimulated by adrenaline, recording the maximum contraction and dT/dt at 12, 36, 60 and 120. When the plateau phase was reached, the vessel was washed with perfusion solution, recording relaxation at 2, 4 and 6. When the base values were reached, Group I underwent a new adrenergic stimulus; and Group II was stimulated with midazolam and then with adrenaline, and the same values were recorded. T test was applied as a statistical analysis when two variables were studied. When studying more than two variables the Anova test was used, supplemented by the Tuckey test.Group I did not show any significant difference between the two stimuli. Group II – the midazolam significantly reduced the maximum contraction induced by adrenaline (83.01 ± 4.11%) (p < 0.01). The dT/dt was reduced at 12 (57.06 ± 8.47%), and also at 36 (70.59 ± 5.26%). There was no significance at 60 and 120 (p < 0.01).The relaxation increased significantly at all measurements – at 2-adrenaline 39.31 ± 9.60%; adrenaline/midazolam: 44.06 ± 9.62% (p < 0.05). At 4-adrenaline: 53.08 ± 8.3%; adrenaline/midazolam: 61.68 ± 8.50% (p < 0.01). At 6-adrenaline: 76.26 ± 5.45%; adrenaline/midazolam: 84.20 ± 7.96% (p < 0.01).Midazolam significantly reduced the maximum contraction obtained by the adrenergic stimulus as well as the dT/dt in the initial phases of contraction. The relaxation speed also increased.     

The reduction of tetrazolium salts by plant mitochondria

Summary Data has been obtained concerning the reduction of tetrazolium salts by mitochondria isolated from Jerusalem artichoke tubers with succinate as the substrate using a direct recording spectrophotometric method of assay. ATP was found to increase the rate of reduction of the tetrazolium salts, this being independent of the effect ATP had on the rate of oxygen uptake. The magnitude of the stimulation by ATP depended on the concentration of tetrazolium salts present and under certain circumstances was suppressed by the addition of azide and cyanide. The sites at which the tetrazolium salts were reduced along the electron transport chain were investigated. The role of ATP has been discussed in relation to the mechanism of tetrazolium reduction.Abbreviations TTC 2,3,5-triphenyl-2,1,3,4-tetrazolium chloride - BT 5,5-diphenyl-3,3-(3,3-dimethoxy-4,4-diphenylene)-ditetrazolium chloride - NT 2,2,5,5-tetraphenyl-3,3-(p-diphenylene)-ditetrazolium chloride - MTT 3-(4,5-dimethyl thiozolyl-2)-2,5-diphenyl tetrazolium bromide - INT 2-(p-iodophenyl)-3-p-dinitrophenyl-3-p-nitrophenyl-5-phenyl tetrazolium chloride - NBT 2,2-dinitrophenyl-5,5-diphenyl-3,3-dimethoxy-4,4-diphenylene)-ditetrazolium chloride - TNBT 2,2-5,5-tetra-p-nitrophenyl-3,3-dimethoxy-4,4-diphenylene) ditetrazolium chloride     

Full 1H NMR assignment of a 24-nucleotide RNA hairpin: Application of the 1H 3D-NOE/2QC experiment

The subject RNA models the binding site for the coat protein of the R17 virus, as well as the ribosome recognition sequence for the R17 replicase gene. With an RNA of this size, overlaps among the sugar protons complicate assignments of the ¹H NMR spectrum. The cross peaks that overlap significantly in 2D-NOE spectra can frequently be resolved by introducing a third, in our approach the double-quantum, frequency axis. In particular the planes in a 3D-NOE/2QC spectrum perpendicular to the 2Q axis are extremely useful, showing a highly informative repeating NOE-2Q pattern. In this experiment substantial J-coupling confers special advantages. This always occurs for geminal pairs (H5/H5 for RNA plus H2/H2 for DNA), as well as for H5/H6, for H3/H4 in sugars with substantial populations of the N-pucker, for H1/H2 for S-puckered sugars, and usually for H2/H3. For the 24-mer RNA hairpin the additional information from the 3D-NOE/2QC spectrum allowed assignment of all of the non-exchangeable protons, eliminating the need for stable-isotope labeling.     

Isolation and structural determination of a novel coenzyme from a methane-oxidizing bacterium

The respiratory quinone composition of the obligate methane-utilizing bacterium Methylomonas rubra was examined. A single lipoquinone was isolated which on examination by thin-layer chromatography cochromatographed with coenzyme Q. Reverse-phase partition and argentation high performance liquid chromatography demonstrated the lipoquinone did not correspond to any known coenzyme Q prenologue. On the basis of mass spectrometry and proton nuclear magnetic resonance spectrometry the novel lipoquinone was shown to correspond to 2,3-dimethoxy-5-methyl-6-(11-methylene-3,7,15, 18, 18, 19, 23-heptamethyltetracosa-2, 6, 14, 19, 22-pentaenyl-)-1,4-benzoquinone.     

The prebiotic chemistry of nucleotides

Diiminosuccinonitrile (DISN), formed by the oxidation of diaminomaleonitrile (DAMN), has been investigated as a potential prebiotic phosphorylating agent. DISN effects the cyclization of 3-adenosine monophosphate to adenosine 2, 3-cyclic phosphate in up to 39% yield. The mechanism of this reaction was investigated. The DISN-mediated phosphorylation of uridine to uridine monophosphate does not proceed efficiently in aqueous solution. The reaction of DISN with uridine-5-phosphate and uridine results in the formation of 2,2-anhydronucleotides and 2,2-anhydronucleosides respectively, and other reaction products resulting from an initial reaction at the 2- and 3-hydroxyl groups. The clay mineral catalysis of the cyclization of adenosine-3-phosphate was investigated using homoionic montmorillonites.     

Polymerization of nucleotide analogues I: Reaction of nucleoside 5′-phosphorimidazolides with 2′-amino-2′-deoxyuridine

Summary 2-Amino-2-deoxyuridine reacts efficiently with nucleoside 5-phosphorimidazolides in aqueous solution. The dinucleoside monophosphate analogues were obtained in yields exceeding 80% under conditions in which little reaction occurs with the natural nucleosides.In a similar way, the 5-phosphorimidazolide of 2-amino-2-deoxyuridine undergoes self-condensation in aqueous solution to give a complex mixture of oligomers.The phosphoramidate bond in the dinucleoside monophosphate analogues is stable for several days at room temperature and pH 7. The mechanisms of their hydrolysis under acidic and alkaline conditions are described.Abbreviations A adenosine - C cytidine - G guanosine - U uridine - T thymidine - U_{N 3} 2-azido-2-deoxyuridine - U_{NH 2} 2-amino-2-deoxyuridine - ImpA adenosine 5-phosphorimidazolide - ImpU uridine 5-phosphorimidazolide - ImpU_{N 3} 2-azido-2-deoxyuridine 5-phosphorimidazolide - ImpU_{NH 2} 2-amino-2-deoxyuridine 5-phosphorimidazolide - pA adenosine 5-phosphate - pU uridine 5-phosphate - pU_{N 3} 2-azido-2-deoxyuridine 5-phosphate - pU_{NH 2} 2-amino-2-deoxyuridine 5-phosphate - UpA uridylyl-[35]-adenosine - UpU uridylyl-[35]-uridine - U_NpA adenylyl-[52]-2-amino-2-deoxy-uridine - U_NpU uridylyl-[52]-2-amino-2-deoxyuridine (pU_N)_n n=2,3,4 [25]-linked oligomers of pU_{NH 2} poly(A) polyadenylic acid - Im imidazole - MeIm l-methylimidazole     

Mineral catalysis of the formation of dimers of 5′-AMP in aqueous solution: The possible role of montmorillonite clays in the prebiotic synthesis of RNA

The reaction of the 5-AMP with water soluble carbodiimide (EDAC) in the presence of Na⁺-montmorillonite 22A results in the formation of 2,5-(pA)₂ (18.9%), 3,5-(pA)₂ (11%), and AppA (4.8%). When poly(U) is used in place of the clay the product yields are 2,5-(pA)₂ (15.5%), 3,5-(pA)₂ (3.7%) and AppA (14.9%). The 3,5-cyclic dinucleotide, 3,5-c(pA)₂, is also formed when poly(U) is used. AppA is the principal reaction product when neither clay nor poly(U) is present in the reaction mixture. Products which contain the phophodiester bond are formed at different ionic strengths, pH and temperatures using Na⁺-montmorillonite. Phosphodiester bond formation was not observed when Cu²⁺-montmorillonite was used or when DISN was used in the place of EDAC. The extent catalysis of phophodiester bond formation varied with the particular clay mineral used. Those Na⁺-clays which bind 5-AMP more strongly are better catalysts. Cu²⁺-montmorillonite, which binds 5-AMP strongly, exhibits no catalytic activity.     

Heteropolynucleotides as templates for non-enzymatic polymerizations

Summary We have studied a number of condensation reactions involving ImpU, ImpT, ImpC, ImpA, ImpG, ImpUpG and ImpCpA as activated nucleotide donors and a variety of homo- and hetero-polynucleotides as templates. We did not obtain any evidence of a template effect with ImpU and ImpT, but observed some condensation of ImpC with GpG on appropriate templates. ImpA and ImpG take part in a number of more or less efficient template-directed reactions, as do ImpUpG and ImpCpA.Our results suggest that, on the primitive Earth, pyrimidine nucleotides could most easily have been incorporated into polymers as constituents of short oligomers, which contained one or more purine nucleotide. The linkage of the product depends strongly on the nature of the substrates; the percentage of the natural 3-5-linkage was, in some cases, less than 10% and, in others, as high as 70%. Wobble-pairing was often very effective in promoting condensations, suggesting that transition mutations would have been very frequent in prebiotic polynucleotide replication.Abbreviations and Conventions U uridine - T thymidine - C cytidine - A adenosine - G guanosine - pN nucleoside-5-phosphate - Np a mixture of 2- and 3-phosphates of a nucleoside - pNp a mixture of the 2-5-diphosphate and 3-5-diphosphate of a nucleoside - N₁ ² pN₂ a 2-5-linked dinucleoside monophosphate - N₁ ³ pN₂ a 3-5-linked dinucleoside monophosphate - N⁵ ppN a pyrophosphate derived from a nucleoside-5-phosphate. ImpN and ImpN₁pN₂ are 5-phosphorimidazolides of nucleosides and 3-5-linked dinucleoside monophosphates, respectively - poly(N) a homopolynucleotide - poly (U₁ C₂ A₄ G₃) a random copolymer derived from a substrate mixture containing U, C, A, G in ratio 1:2:4:3 - ODU optical density units measured at 260 nm     

Degradation of the diarylpropane lignin model compound 1-(3′,4′-diethoxyphenyl)-1,3-dihydroxy-2-(4′'-methoxyphenyl)-propane and derivatives by the basidiomycete Phanerochaete chrysosporium

The white rot basidiomycete Phanerochaete chrysosporium metabolized 1-(3,4-diethoxyphenyl)-1,3(dihydroxy)-2-(4'-methoxyphenyl)-propane (XII) in low nitrogen stationary cultures, conditions under which the ligninolytic enzyme system is expressed. 3,4-Diethoxybenzyl alcohol (IV), 1,2(dihydroxy)-1-(4-methoxyphenyl)ethane (XX) and anisyl alcohol were isolated as metabolic products indicating an initial , bond cleavage of this dimer. Exogenously added XX was rapidly converted to anisyl alcohol, indicating that XX is an intermediate in the metabolism of XII. Fungal cleavage of the , bond of 1-(3-4-diethoxyphenyl)-1-(hydroxy)-2-(4'-methoxyphenyl)ethane (XI) also occurred, indicating that a hydroxymethyl group is not a prerequisite for this reaction. P. chrysosporium also metabolized 1-(4-ethoxy-3-methoxyphenyl)-2,2(dihydroxy)-2-(4'-methoxyphenyl)propane-1-ol (XIII). The major products of the degradation of this triol included 4-ethoxy-3-methoxybenzyl alcohol (III) and 2-hydroxy-1-(4-methoxyphenyl)-1-oxoethane (XXI). The nature of the products formed indicates that this triol is also cleaved directly at the , bond. The significant difference in the nature of the products formed from the diaryl propane (XII) and the triol (XIII), however, suggests that XIII is not an intermediate in the major pathway for the degradation of XII. Metabolites were identified after comparison with chemically synthesized standards by GLC-mass spectrometry.Abbreviations GLC Gas liquid chromatography - TMSi trimethylsilyl - TLC thin layer chromatography - MS mass spectrometry     

5′-Nucleotidase

Summary To get more insight in the function of 5-nucleotidase catabolic and anabholic processes were investigated in which 5-nucleotides are involved. The catabolism of adenosine-5-monophosphate was studied by investigating the reaction products obtained after incubation of homogenates of several organs of rat and mouse with adenosine-5-monophosphate and with adenosine. Two experimental tumours of the mouse were investigated in the same way. It was found that in tissues containing a high activity of 5-nucleotidase other enzymes involved in the catabolism of 5-nucleotides, such as nucleosidase, adenosine deaminase and adenosine-5-monophosphate deaminase could also be demonstrated.The anabolic processes in which 5-nucleotides are involved had been studied by investigating the incorporation of tritium-labeled thymidine in several tissues of the mouse. It appeared that in cells showing a high 5-nucleotidase activity no incorporation of radioactive thymidine could be found, while in cells showing incorporation of thymidine enzyme activity could not be demonstrated.A discussion is given about the possible role of 5-nucleotidase in the control of nucleic acid biosynthesis and in the catabolism of nucleic acids.Abbreviations used DNA deoxyribonucleic acid - RNA ribonucleic acid - AMP Adenosine-5-monophosphate - ADP Adenosine-5-diphosphate - ATP Adenosine-5-triphosphate - IMP Inosine-5-monophosphate - GMP Guanosine-5-monophosphate - GDP Guanosine-5-diphosphate - GTP guanosine-5-triphosphate - CMP Cytidine-5-monophosphate - CDP Cytidine-5-diphosphate - CTP Cytidine-5-triphosphate - UMP Uridine-5-monophosphate - UDP Uridine-5-diphosphate - UTP Uridine-5-triphosphate - TMP Thymidine-5-monophosphate - TDP Thymidine-5-diphosphate - TTP Thymidine-5-triphosphate - Ado Adenosine - Ad Adenine - Ino Inosine - Hypox Hypoxanthine - Xanth Xanthine - Xantho Xanthosine - Guano Guanosine - Gua Guanine - Ura Uracil - U Uridine - Cyt Cytidine - Cyto Cytosine - Thym Thymidine The corresponding deoxy-compounds have been indicated with the prefix d for instance dCMP, deoxycytidine-5-monophosphate.     

Indigoidine and other bacterial pigments related to 3,3′-bipyridyl

Zusammenfassung Die extracelluläre Abscheidung eines unlöslichen blauen Pigments (Indigoidin) wurde zuerst bei Pseudomonas indigofera beobachtet. Historisch wird auf die verschiedenen Benennungen dieses Bakteriums eingegangen. Beschrieben wird die Darstellung blauer Farbstoffe aus Kulturen verschiedener Bakterien. Die von Corynebacterium insidiosum, Arthrobacter atrocyaneus und Arthrobacter polychromogenes gebildeten Pigmente sind identisch mit Indigoidin von P. indigofera. Die Identität wird bewiesen durch physikalische und chemische Vergleiche der Pigmente und ihrer Derivate. Der Name Indigoidin, der früher nur für das Pigment von P. indigofera verwendet wurde, wird nun unabhängig von der Herkunft des Pigments benützt.Indigoidin (I), C₁₀H₈N₄O₄, ist 5,5-Diamino-4,4-dihydroxy-3,3-diazadiphenochinon-(2,2). Durch Erhitzen mit 6 n HCl entsteht daraus ein Hydrolyseprodukt (III), C₁₀H₆N₂O₆, das als 4,5,4,5-Tetrahydroxy-3,3-diazadiphenochinon-(2,2) erkannt wurde. Dieses Hydrolyseprodukt (III) bildet ein Monokaliumsalz (VII), das identisch ist mit dem grünen Pigment, das Arthrobacter crystallopoietes bei Zusatz von Pyridon-(2) bildet. Über Synthesen des Indigoidins (I) und seines Hydrolyseprodukts (III), die von 3,3-Bipyridyl, von Citrazinsäure oder 5-Amino-pyridon-(2) ausgehen, wird an anderer Stelle berichtet.Beschrieben wird die Darstellung folgender Indigoidin-Derivate: 5,5-Diacetamino-4,4-dihydroxy-3,3-diazadiphenochinon-(2,2) (II), C₁₄H₁₂N₄O₆; 4,4-Dihydroxy-5,5-diacetoxy-3,3-diazadiphenochinon-(2,2) (IV), C₁₄H₁₀N₂O₈; 2,5,6,2.5.6-Hexaacetoxy-3,3-bipyridyl (VI), C₂₂H₂₀N₂O₁₂ und 4,4-Dihydroxy-5,5-dimethoxy-3,3-diazadiphenochinon-(2,2) (V), C₁₂H₁₀N₂O₆.     

Adenosine-5′-phosphosulfate (APS) as sulfate donor for assimilatory sulfate reduction in Rhodospirillum rubrum

Crude extracts of Rhodospirillum rubrum catalyzed the formation of acid-volatile radioactivity from (³⁵S) sulfate, (³⁵S) adenosine-5-phosphosulfate, and (³⁵S) 3-phosphoadenosine-5-phosphosulfate. An enzyme fraction similar to APS-sulfotransferases from plant sources was purified 228-fold from Rhodospirillum rubrum. It is suggested here that this enzyme is specific for adenosine-5-phosphosulfate, because the purified enzyme fraction metabolized adenosine-5-phosphosulfate, however, only at a rate of 1/10 of that with adenosine-5-phosphosulfate. Further, the reaction with 3-phosphoadenosine-5-phosphosulfate was inhibited with 3-phosphoadenosine-5-phosphate whereas this nucleotide had no effect on the reaction with adenosine-5-phosphosulfate. For this activity with adenosine-5-phosphosulfate the name APS-sulfotransferase is suggested. This APS-sulfotransferase needs thiols for activity; good rates were obtained with either dithioerythritol or reduced glutathione; other thiols like cysteine, 2-3-dimercaptopropanol or mercaptoethanol are less effective. The electron donor methylviologen did not catalyze this reaction. The pH-optimum was about 9.0; the apparent K _m for adenosine-5-phosphosulfate was determined to be 0.05 mM with this so far purified enzyme fraction. Enzyme activity was increased with K₂SO₄ and Na₂SO₄ and was inhibited by 5-AMP. These properties are similar to assimilatory APS-sulfotransferases from spinach and Chlorella.Abbreviations APS adenosine-5-phosphosulfate - PAPS 3-phosphoadenosine-5-phosphosulfate - 5-AMP adenosine-5-monophosphate - 3-AMP adenosine-3-monophosphate - 3-5-ADP 3-phosphoadenosine-5-phosphate (PAP) - DTE dithiorythritol - GSH reduced glutathione - BAL 2-3-dimercaptopropanol     

Effects of cyclic nucleotides on morphogenesis inNostoc muscorum

The filamentous cyanophyteNostoc muscorum A grew aseriately in light in a mineral salts (sugar-free) culture medium supplemented with adenosine 3:5-cyclic-monophosphate or N⁶, O²-dibutyryl adenosine 3:5-cyclic-monophosphate (1 mM). The aseriate morphology thus formed in the light on the 10th day following inoculation was similar to that formed in the dark after 20–30 days growth in cAMP-free medium containing glucose or sucrose. Inoculum previously grown in sucrose- or glucose-containing medium displayed aseriate morphology with lesser proliferation of coccoid cells as compared to inoculum grown in the absence of glucose or sucrose. cGMP, ADP, AMP and inhibitors of phosphodiesterase (theophylline and caffeine) did not have any effect on the persistence of aseriate morphology. However they stimulated cell division at the aseriate stage and delayed the release of hormogonia.Abbreviations cAMP adenosine 3:5-cyclic-monophosphate - db cAMP N⁶, O²-dibutyryl adenosine 3:5-cyclic-monophosphate - cGMP guanosine 3:5-cyclic-monophosphate - ATP adenosine 5-triphosphate - ADP adenosine5-diphosphate - AMP adenosine 5-monophosphate     

Determination of internal dynamics of deoxyriboses in the DNA hexamer d(CGTACG)2 by 1H NMR

The conformations and internal dynamics of the deoxyriboses of d(CGTACG)₂ have been determined by NMR measurements at 15°C. The conformations of the sugars were determined using coupling constants and time-dependent NOE measurements. The J-splitting patterns of the H1, H2 and H2 resonances show that the sugars exist as mixtures of conformations near C2 endo (south) and C3 endo (north). The population of the south conformation was larger for the purines than for the pyrimidines. The overall tumbling time of the molecule in ²H₂O was determined from measurements of the cross relaxation rate constant for the H6-H5 vectors of the two cytosine residues. Order parameters were determined for the H1-H2, H2-H2 and H2-H3 vectors from measurements of cross relaxation rate constants, making use of multi-spin analysis of the NOE build up rates. These order parameters are weakly dependent of the base sequence, and except for the terminal Cyt 1 residue, the H2-H2 and H2-H3 vectors are near unity, indicating the absence of rapid pseudorotation on the nanosecond time scale. However, the order parameter for the H1-H2 vector is significantly smaller than expected for rapid pseudorotation indicating the presence of other motions of the sugars. This motion must be about an effective axis parallel to the H2-H vector, and to occur with an angular fluctuation of about 30°.The results show that to obtain highly refined structures for nucleic acids by NMR the effects of spin diffusion and motional averaging cannot be ignored.Some of this work was presented as a poster at the 30^th Experimental NMR Conference at Asilomar, California 1989     

Determination of the backbone torsion angle ɛ in nucleic acids

Summary The multiplet structure of cross peaks in double-quantum-filtered COSY NMR spectra is analysed for those resonances that include passive heteronuclear couplings. Interestingly, the cross peak involving the sugar-ring protons H2 and H3 in nucleic acids display an E.COSY-type appearance exclusively when the backbone torsion angle (C4-C3-O3-P) adopts a gauche(-) conformation. This observation allows an unambiguous analysis of the conformation around , without the knowledge of ³J_cp coupling constants.     

Nyctohemeral rhythmicity of type II thyroxine 5′-deiodinase activity in the pineal gland but not in the Harderian gland of the Swiss mouse

Type II thyroxine 5-deiodinase (5-D) activity in both pineal and Harderian glands of the Swiss mouse was studied. Pineal 5-D activity exhibited a nyctohemeral profile with a maximal peak value at 05.00 h, which coincides with that for pineal melatonin production. However, no rhythm of 5-D activity in the Harderian gland could be found. In pineal gland, light at night inhibited the nocturnal increase in 5-D activity, while isoproterenol, a -adrenergic agonist, could not stimulate the enzyme. In the Harderian gland, neither darkness, nor light at night, or isoproterenol were capable of modifying basal values of 5-D activity.