A novel PH-CT-COSY methodology for measuring JPH coupling constants in unlabeled nucleic acids. Application to HIV-2 TAR RNA

A quantitative analysis of J_PH scalar couplings in nucleic acids is difficult due to small couplings to phosphorus, the extreme overlap of the sugar protons and the fast relaxation of the spins involved in the magnetization transfer. Here we present a new methodology that relies on heteronuclear Constant Time Correlation Spectroscopy (CT-COSY). The three vicinal ³J_PH3, ³J_PH5 and ³J_PH5 scalar couplings can be obtained by monitoring the intensity decay of the P_i-H3_{i – 1} peak as a function of the constant time T in a 2D correlation map. The advantage of the new method resides in the possibility of measuring the two ³J_PH5 and ³J_PH5 scalar couplings even in the presence of overlapped H5/H5 resonances, since the quantitative information is extracted from the intensity decay of the P-H3 peak. Moreover, the relaxation of the H3 proton is considerably slower than that of the H5/H5 geminal protons and the commonly populated conformations of the phosphate backbone are associated with large ³J_PH3 couplings and relatively small ³J_{PH5 / H5}. These two facts lead to optimal signal-to-noise ratio for the P-H3 correlation compared to the P-H5/H5 correlation.The heteronuclear CT-COSY experiment is suitable for oligonucleotides in the 10–15 kDa molecular mass range and has been applied to the 30mer HIV-2 TAR RNA. The methodology presented here can be used to measure P-H dipolar couplings (D_PH) as well. We will present qualitative results for the measurement of P-H_base and P-H2 dipolar couplings in the HIV-2 TAR RNA and will discuss the reasons that so far precluded the quantification of the D_PHs for the 30mer RNA.     

Enzymatic semisynthesis of aprotinin homologues mutated in P′ positions

The replacement of amino acids in the P₁ and P₂ position of aprotinin, the bovine pancreatic trypsin inhibitor, is described. Using the modified inhibitor as starting material, with the hydrolyzed reactive-site peptide bond Lys¹⁵-Ala¹⁶, the residues P₁ (Ala¹⁶) and P₂ (Arg¹⁷) were split off by the action of aminopeptidase K. Incorporation of suitable dipeptides containing a basic residue (Lys or Arg) in the C-terminal position was carried out in a one pot reaction involving trypsin-catalyzed coupling. In this way, the native fragment Ala¹⁶-Arg¹⁷ was reintroduced and also replaced by Gly-Arg, Ala-Lys, and Leu-Arg yielding intact inhibitor molecules. The mechanism for incorporation of dipeptides was investigated by treating the aprotinin derivative with the Arg¹⁷-Ile¹⁸ peptide bond hydrolyzed with trypsin under proteosynthetic conditions. We established that only inhibitor molecules cleaved between Lys¹⁵ and Xaa¹⁶ are intermediates leading to the desired products. The inhibitory properties of the new aprotinin homologues were tested, and the significance of the P₁ residue for the inhibition of trypsin, kallikrein, and chymotrypsin was deduced.     

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     

A fluorometric assay of ceramide glycanase with 4-methylumbelliferyl β-d-lactoside derivatives

4-Methylumbelliferyl 6-O-benzyl--d-lactoside (6Bn-MU-Lac) and some related compounds were synthesizedvia different selective reactions including phase-transfer glycosylation. Their suitability as substrates for a fluorometric assay of ceramide glycanase (CGase) was evaluated. Among others, the 6Bn-MU-Lac, which is resistant to exogalactosidase, was found to be a suitable substrate for routine assay of the CGase activity. For American leech CGase, theK _m value is 0.232 mM at pH 5. Abbreviations: CGase, ceramide glycanase; Gal, galactose; Glc, Glucose; Lac, lactose; MU, 4-methylumbelliferone; MU-Lac, 4-methylumbelliferyl -d-lactoside; bBn-Lac, 6-O-benzyl-lactose; 6Bn-MU-Lac, 4-methylumbelliferyl 6-Obenzyl--d-lactoside; 46Bd-MU-Lac, 4-methylumbelliferyl 4,6-O-benzylidene--d-lactoside; MU-Cel, 4-methylumbellifery -d-cellobioside; 46Bd-MU-Cel, 4-methylumbelliferyl 4,6-O-benzylidene--d-cellobioside; TLC, thin layer chromatography;¹H-NMR, proton nuclear magnetic resonance; GSL, glycosphingolipids; CSA, 10-camphorsulfonic acid. See Scheme 1 for chemical structures.     

Oxidation of cysteine to cystine by membrane fractions of Chlorella fusca

Isolated membrane fractions of Chlorella fusca 211-8b obtained by french-press treatment and sonication catalyzed the oxidation of l-cysteine to l-cystine. The pH-optimum of this reaction was determined to be around 8–8.5 and a stoichiometry of 4 SH-groups oxidized for one O₂ consumed was obtained. This thiol-oxidation system was specific for D-and l-cysteine; Dl-homocysteine and cysteamine were oxidized at about half the rate whereas all other thiols tested including glutathione, mercaptoethanol, mercaptopropionic acid and dithioerythritol were not oxidized by these membrane fractions. The apparent K_m for l-cysteine was determined as 3.3 mmol l^-1. Rates of 200 mol cysteine oxidized mg^-1 chlorophyll h^-1 were normally obtained. Extremely high rates of oxygen uptake were measured using l-cysteine methyl ester and l-cysteine ethyl ester. This thioloxidation system was not inhibited by mitochondrial electron-transport inhibitors such as rotenone or antimycin A, nor by the chloroplast electron-transport inhibitors 2,5-dibromothymochinone and 2,4-dinitrophenylether of iodonitrothymol. The cysteine oxidation catalyzed by C. fusca membranes was inhibited, however, by salicylhydroxamic acid, o-phenanthrolin, N,N-disalicyliden-1,3-diaminopropane 5,5-disulfonic acid, ethylenediaminetetraacetic acid, high KCN levels and by the buffers, N-[2-hydroxyl-1,1-bis(hydroxymethyl) ethyl] glycine and phosphate. This cysteine-oxidation system seems to function as a counterpart of thioredoxin-mediated light activation of enzymes, allowing reduced thiol groups to be oxidized again by O₂ (dark inactivation).Abbreviation DTNB 5,5-dithio-bis(-2-nitrobenzoic acid). Ellmann reagent     

Comparison of Some Photosynthetic Characters Between Two Hybrid Rice Combinations Differing in Yield Potential

Photosynthetic characteristics of two hybrid rice combinations, Peiai 64S/E32 and Shanyou 63, were compared at the panicle differentiation stage. As compared with Shanyou 63, the new combination Peiai 64S/E32 showed a significantly higher net photosynthetic rate (P _N), apparent quantum yield of carbon assimilation (_c), carboxylation efficiency (CE), and photorespiratory rate (R _P) as well as leaf chlorophyll content, but a significantly lower dark respiration rate (R _D) and compensation irradiance (I _c). It also showed a slightly higher photochemical efficiency (F_v/F_m and F/F_m) of photosystem 2, a lower non-photochemical quenching (q_N), and a similar CO₂ compensation concentration () as compared to Shanyou 63.     

Inactivation of DNA polymerases by adenosine 2′,3′-riboepoxide 5′-triphosphate allows estimation of the primers affinity

Template-primer dependent inactivation of human DNA polymerase and Klenow fragment of E. coli DNA polymerase I by adenosine 2,3-riboepoxide 5-triphosphate was used for quantitative analysis of the K_d values for oligonucleotide primers of different length. The K_d values are smaller by a factor of 2.5 than the K_m values for the same primers determined in the reaction of DNA polymerization in the case of DNA polymerase . The K_d and K_m values are nearly the same for Klenow fragment. Such approach to the determination of K_m/K_d ratio can likely be used for detailed quantitative analysis of DNA polymerases.Abbreviations epATP adenosine 2,3-riboepoxide 5-triphosphate - KF Klenow fragment of E. coli DNA polymerase I - Pol I E. coli DNA polymerase I - Pol human placenta DNA polymerase      

Microbiological synthesis of 5-ethyl- and (E)-5-(2-bromovinyl)-2′-deoxyuridine

Summary The title compounds were prepared by an enzymatic transdeoxyribosylation from 2 dGuo or 2 dThd to the respective heterocyclic bases, 5-ethyluracil and (E)-5-(2-bromovinyl)uracil, using the whole bacterial cells ofEscherichia coli as a biocatalyst.     

The role ofc-type cytochromes in the terminal respiratory chain of the methylotrophic bacteriumMethylophilus methylotrophus

Whole cells of the methylotrophic bacteriumMethylophilus methylotrophus cultured under methanol-limited conditions contain approximately equal amounts of two majorc-type cytochromes,c _H andc _L. Virtually all of the cytochromec _H, and over one-third of the cytochromec _L, are loosely attached to the periplasmic surface of the respiratory membrane whence they can be released by sonication or by washing cells in ethylenediaminetetraacetate (EDTA). The latter causes inhibition of methanol oxidase activity and stimulation of ascorbate-N,N,N,N-tetramethyl-p-phenylenediamine (TMPD) oxidase activity, neither of which effects are reversible by divalent metal ions. Kinetic analyses indicate that ascorbate-TMPD is oxidised via two routes, viz. a slow low-affinity pathway involving loosely membrane-boundc-type cytochromes plus cytochrome oxidaseaa ₃, and a faster higher-affinity pathway involving the firmly membrane-bound cytochrome oxidasec _L o complex; the former route predominates in the presence of divalent metal ions, and the latter route after exposure to EDTA. These and other results are discussed in terms of the spatial organisation of the terminal respiratory chain, and of the role ofc-type cytochromes in the oxidation of methanol and ascorbate-TMPD.Abbreviations EDTA Enthylenediaminetetraacetate - PMS Phenazinemethosulphate - TMPD N,N,N,N-tetramethyl-p-phenylenediamine - SDS Sodium dodecylsulphate - I₅₀ Concentration of inhibitor required to give 50% inhibition of enzyme activity - PQQ Pyrroloquinoline quinone     

Restoration of the wild-type locus in an RuBP carboxylase/oxygenase mutant of Synechocystis PCC 6803 via targeted gene recombination.

Summary The interaction between homologous DNA sequences, distant from each other in the chromosome, was examined in the cyanobacterium Synechocystis PCC 6803. Most of the rbcL gene encoding the large subunit of ribulose bisphosphate carboxylase/oxygenase (Rubisco) was duplicated in the genome by a targeted insertion of a 3-truncated gene copy into the psbA-I locus. Both rbcL genes, in the psbA-I region and at the rbc locus, were non-functional; The former due to the 3 truncation, and the latter due to a deletion in the 5-region (creating a 5 truncation) and a mutation associated with an insertion of the Rhodospirillum rubrum rbc gene, yielding a high-CO₂-requiring mutant (cyanorubrum). The 3 and the 5 truncated rbcL genes were linked to chloramphenicol and kanamycin resistance markers, respectively. Decreasing the kanamycin selective pressure concomitantly with exposure of the double resistance mutant to air, resulted in air-growing colonies. Analysis of their genomes, Rubisco proteins, and their ultrastructure revealed: 1) Reconstitution of a full-length cyanobacterial rbcL gene at the rbc locus; 2) simultaneous synthesis of the cyanobacterial (L₈S₈) and R. rubrum L₂) enzymes in meroploids containing both mutated and reconstituted rbcL genes; 3) reappearance of carboxysomes. Our results indicate extensive recombinatorial interactions between the homologous sequences at both loci leading to reconstitution of the cyanobacterial rbcL gene.     

Conformational dynamics of the anticodon loop in yeast tRNAPhe as sensed by the fluorescence of wybutine

Conformational and dynamic properties of the anticodon loop of yeast tRNA^Phe were investigated by analyzing the time resolved fluorescence of wybutine serving as a local structural probe adjacent to the anticodon GmAA on its 3 side. The influence of Mg²⁺, important for stabilizing the tertiary structure of tRNA, and of the complementary anticodon s²UUC of E. coli tRNA ₂ ^Glu were investigated.Fluorescence lifetimes and anisotropies were measured with ps time resolution using time correlated single photon counting and a mode locked synchronously pumped and frequency doubled dye laser as excitation source. From the analysis of lifetimes () and rotational relaxation times ( _R ) we conclude that wybutine occurs in various structural states: (i) one stacked conformation where the base has no free mobility and the only rotational motion reflects the mobility of the whole tRNA molecule (=6 ns, _R =19 ns), (ii) an unstacked conformation where the base can freely rotate (=100 ps, _R = 370 ps) and (iii) an intermediary state (=2 ns, _R = 1.6 ns).Under biological conditions, i. e. in the presence of Mg²⁺ and neutral salts, wybutine is found in a stacked and immobile state which is consistent with the crystallographic picture. In the presence of the complementary codon however, as exemplified by the E. coli-tRNA ₂ ^Glu anticodon, our analysis indicates that the codon-anticodon complex exists in an equilibrium of structural states with different rotational mobility of wybutine. The conformation with wybutine freely mobile is the predominant one and suggests that this conformation of the codon-anticodon structure differs from the canonical 3–5 stack.     

Interaction between photosynthetic and respiratory electron-transfer chains in the membranes of Anabaena variabilis

The rate of CO₂- and p-benzoquione-dependent photosynthetic O₂ evolution by Anabaena variabilis cells remained unaltered and the rate of O₂ uptake observed after switching off the light (endogenous respiration) was enhanced by a factor of 6–8 when the O₂ concentration was increased from 200 to 400 M. Photosystem-I-linked O₂ uptake and respiration of the cells incubated with ascorbate and N,N,NN-tetramethyl-p-phenylenediamine was not appreciable influenced by the O₂ concentration. 2-Iodo-6-isopropyl-3-methyl-2,4,4-trinitrodiphenyl ether, blocking electron transfer at the plastoquinone level, suppressed O₂ evolution and had no influence on endogenous respiration. 2-n-Heptyl-4-hydroxyquinoline-N-oxide, an inhibitor of electron transfer between photosystems II and I, as well as the cytochrome-oxidase inhibitors N ₃ ^- , CN^- and NH₂OH, caused a 35–50% retardation of endogenous respiration and blocked photosynthetic O₂ evolution. The molar ratio of cytochromes b₆, f, c-553, aa₃ and photosystem-I reaction centers in the isolated membranes equalled approx. 2:1:2:0.7:2. It is inferred that endogenous respiration of A. variabilis cells is inhibited by the light-induced electron flow through both photosystems at the level of the plastoquinone-plastocyanin-oxidoreductase complex.Abbreviations DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DNP-INT 2-iodo-6-isopropyl-3-methyl-2,4,4-trinitrodiphenyl ether - Hepes 4-(2-hydroxyethyl)-1-piperazine ethansulfonic acid - TMPD N,N,NN-tetramethyl-p-phenylenediamine     

Geometry of the dry-state oligomerization of 2′,3′-cyclic phosphates

Summary Evaporation of a solution of thymidine plus either theexo or theendo diastereomer of uridine cyclic 2,3-O, O-phosphorothioate (U > p(S) in 1,2-diaminoethane hydrochloride buffer gave the 2,5 and 3,5 isomers of (P-thio) uridylylthymidine (Up(S)dT) in a ratio of 1:2 with a combined yield of about 20%. These isomers were re-converted to U > p(S) and dT by a reaction that is known to proceed by an in-line mechanism. Both the 2,5 and 3,5 isomers gave as product the same diastereomer of U > p(S) that had been used originally in their formation. These dry-state prebiotic reactions (Verlander, Lohrmann, and Orgel 1973) are thus shown to be stereospecific, and both the 2,5 and 3,5 internucleotide bonds are formed by an in-line mechanism.Abbreviations DAE 1,2-diaminoethane - HPLC high pressure liquid chromatography - RNase bovine pancreatic ribonuclease A, EC 3.1.4.22 - TEAB triethylammonium bicarbonate - tris tris(hydroxymethyl)aminomethane - UMP(S) uridine monophosphorothioate - U > p uridine cyclic 2,3-phosphate - U > p(S) uridine cyclic 2,3-O, O-phosphorothioate - Up(S)dT (P-thio)uridylylthymidine - U2p(R_p-S)5dT (P-thio)uridylylthymidine with theR configuration at phosphorous, and a 2,5 internucleotide linkage     

Two-dimensional 1H NMR study of two cyclobutane type photodimers of thymidylyl-(3′→5′)-thymidine

2D NMR spectroscopy and J coupling constant analysis are applied to resolve the structure of two photoproducts of thymidylyl-(35)-thymidine. These products are cyclobutane type thymine dimers possessing the cis-syn (the predominant one) and trans-syn geometry. The cis-syn is formed in an ANTI-ANTI conformation about the N-glycosyl linkages and resembles the normal base-stacked configuration. The glycosidic conformation in solution of the 5 terminal fragment differs from the crystal in which the less common SYN conformation is observed. In this isomer only the sugar pucker of the 3 terminal fragment is changed substantially with respect to the dinucleotide. The trans-syn isomer is formed in a SYN-ANTI glycosidic conformation. In this isomer the sugar puckers of both deoxyribose rings are affected and a preference for a pure 2-endo conformation is observed.Abbreviations dTpdT 2-deoxythymidylyl-(35)-2-deoxythymidine - dTp[]dT cyclobutane type photodimers of dTpdT - dTp- and dTp[]- their 5' terminal fragments (fragment A) - -pdT and-[]pdT their 3 terminal fragments (fragment B) - RP-HPLC reversed-phase high-performance liquid chromatography - COSY two-dimensional correlated spectroscopy - 2D NOE two-dimensional nuclear Overhauser spectroscopy     

The formation of peptides from the 2′(3′)-glycyl ester of a nucleotide

Summary We have synthesized 2(3)-O-(glycyl)-adenosine-5-(O-methylphos-phate), an analogue of the 3-terminus of aminoacylated tRNA. A 0.4M solution of this compound maintained at pH 8.2, yields 5.5% of diglycine and 11.5% of diketopiperazine, in addition to the hydrolysis products glycine and adenosine-5-(O-methylphosphate). Under the same conditions, glycine ethyl ester reacts much more slowly, but ultimately gives similar yields of diglycine and diketopiperazine.The aminolysis of 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate) by free glycine is relatively inefficient, but serine reacts 20 times more rapidly and yields up to 50% of N-glycylserine. The prebiotic significance of these reactions is discussed.Abbreviations MepA adenosine-5-(O-methylphosphate) - MepA-gly 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate) - MepA-bis-gly 2,3-O-(bis-glycyl)-adenosine-5-(O-methylphosphate) - DKP diketopiperazine - gly Et glycine ethyl ester - gly-ser N-glycylserine - O-gly-ser O-glycylserine - O-(gly)-gly-ser O-(glycyl)-glycylserine - Boc-gly N-tert-butyloxycarbonylglycine - MepA-Boc-gly 2(3)-O-(Boc-glycyl)-adenosine-5-(O-methylphosphate) - MepA-bis-Boc-gly 2,3-O-(bis-Boc-glycyl)-adenosine-5(O-methylphosphate) - (gly)₂ diglycine - (gly)₃ triglycine     

Cyclic AMP regulation of glucose transport in germinating Pilobolus longipes spores

Pilobolus longipes spores were activated by either glucose or 6-deoxyglucose. Glucose-induced spore activation was previously shown to follow an increase in intracellular cyclic AMP. Concurrent with glucose-induced spore activation, were shifts in 6-deoxyglucose transport kinetics towards higher V _max and K _m values. Cyclic AMP derivatives also caused spore activation and similar changes in the kinetic parameters of 6-deoxyglucose transport. The time course of activation was paralleled by changes in transport activity. Inhibition of phosphodiesterase alone did not cause activation or induce changes in transport activity, but in combination with sub-optimal levels of either 6-deoxyglucose or cAMP derivatives, it amplified the germination signals to produce large increases in both spore activation and 6-deoxyglucose transport activity. These results support the conclusion that glucose transport in germinating spores is regulated by cAMP.Abbreviations IBMX 3-isobutyl-1-methylxanthine; monobutyryl cyclic AMP - N⁶ monobutyryladenosine 3:5-cyclic monophosphate - 8-bromo cyclic AMP 8-bromoadenosine 3:5-cyclic monophosphate     

Nucleotide sequence of the rrnB 16S ribosomal RNA gene from Mycoplasma capricolum

Summary The nucleotide sequences of the rrnB 16S ribosomal RNA gene and its 5-and 3-flanking regions from Mycoplasma capricolum have been determined. The coding sequence is 1521 base pairs long, being 21 base pairs shorter than that of the Scherichia coli 16S rRNA gene. The 16S rRNA sequence of M. capricolum reveals 74% and 76% identity with that of E. coli and Anacystis nidulans, respectively. The secondary structure model constructed from the M. capricolum 16S rRNA.gene sequence resembles that proposed for E. coli 16S rRNA. A large stem structure can be constructed between the 5- and 3-flanking sequences of the 16S rRNA gene. The flanking regions are extremely rich in AT.     

Non-PTS uptake and subsequent metabolism of glucose in Pediococcus halophilus as demonstrated with a double mutant defective in phosphoenolpyruvate:mannose phosphotransferase system and in phosphofructokinase

Pediococcus halophilus possesses phosphoenolpyruvate:mannose phosphotransferase system (man:PTS) as a main glucose transporter. A man:PTS defective (man:PTS^d) strain X-160 could, however, utilize glucose. A possible glucose-transport mechanism other than PTS was studied with the strain X-160 and its derivative, man:PTS^d phosphofructokinase defective (PFK^–) strain M-13. Glucose uptake by X-160 at pH 5.5 was inhibited by any of carbonylcyanide m-chlorophenylhydrazone, nigericin, N,N-dicyclohexylcarbodiimide, or iodoacetic acid. The double mutant M-13 could still transport glucose and accumulated intracellularly a large amount of hexose-phosphates (ca. 8 mM glucose 6-phosphate and ca. 2 mM fructose 6-phosphate). Protonophores also inhibited the glucose transport at pH 5.5, as determined by the amounts of accumulated hexose-phosphates (< 4 mM). These showed involvement of proton motive force (P) in the non-PTS glucose transport. It was concluded that the non-PTS glucose transporter operated in concert with hexokinase or glucokinase for the metabolism of glucose in the man:PTS^d strain.Abbreviations BM basal medium - BM-G basal medium containing glucose - CM complex medium - man:PTS phosphoenolpyruvate:mannose phosphotransferase system - CCCP carbonylcyanide m-chlorophenylhydrazone - DCCD N,N-dicyclohexyl carbodiimide - P proton motive force - pH transmembrane pH gradient - transmembrane electrical potential difference - MNNG N-methyl-N-nitro-N-nitrosoguanidine - PIPES piperazine-N,N-bis(-ethanesulfonic acid) - MES 4-morpholineethanesulfonic acid - G-6-P glucose 6-phosphate - F-6-P fructose 6-phosphate - FDP fructose 1,6-bisphosphate - EMP Embden-Meyerhof-Parnas pathway - PFK phosphofructokinase - GK glucokinase - HK hexokinase - IAA iodoacetic acid - II^man enzyme II component of man:PTS     

Elucidation of Structure of Lipid A from the Marine Gram-Negative Bacterium Pseudoalteromonas haloplanktis ATCC 14393T

The chemical structure of lipid A, from the marine -proteobacterium Pseudoalteromonas haloplanktis 14393, a main product of lipopolysaccharide hydrolysis (1% AcOH), was determined using chemical methods and NMR spectroscopy. The lipid A was shown to be -1,6-glucosaminobiose 1,4-diphosphate acylated with two (R)-3-hydroxyalkanoic acid residues at C3 and C3 and amidated with one (R)-3-hydroxydodecanoyl and one (R)-3-dodecanoyloxydodecanoyl residue at N2 and N2, respectively.     

The catalysis of nucleotide polymerization by compounds of divalent lead

Summary Soluble lead salts and a number of lead-containing minerals catalyze the formation of oligonucleotides from nucleoside 5-phosphorimidazolides. The effectiveness of lead compounds correlates strongly with their solubility. Under optimal conditions we were able to obtain 18% of pentamer and higher oligomers from ImpA. Reactions involving ImpU gave smaller yields.Abbreviations A adenosine - U uridine - Im imidazole - MeIm 1-methyl-imidazole - EDTA ethylenediaminetetraacetic acid - pA adenosine 5-phosphate - pU uridine 5-phosphate - Ap adenosine cyclic 2:3-phosphate - ATP adenosine 5-triphosphate - AppA P₁,P₂-diadenosine 5-diphosphate - pNp (N = A,U) nucleotide 2(3), 5-diphosphate - ImpA adenosine 5-phosphoreimidazolide - ImpU uridine 5-phosphorimidazolide - A ²pA adenylyl-[25]-adenosine - A ³pA adenylyl-[35]-adenosine - pA ²pA 5-phospho-adenylyl-[25]-adenosine - pA ³pA 5-phospho-adenylyl-[35]-adenosine - pUpU 5-phospho-uridylyl-uridine - pApU 5-phospho-adenylyl-uridine - pUpA 5-phospho-uridylyladenine - (pA)n (n, 2,3,4,) oligoadenylates with 5 terminal phosphate - ImpApA 5-phosphorimidazolide of adenylyl adenosine - (pA) ₅₊ pentamer and higher oligoadenylates with 5 terminal phosphate - (Ap)nA (n = 2,3,4) oligoadenylates without terminal phosphates In the following we do not specify the nature of the internucleotide linkageIn the following we do not specify the nature of the internucleotide linkage