首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 31 毫秒
1.
Myobacterium avium LM1 was exposed to concentrations of 5-fluorouracil (5FU) that ranged from 0 to 100 g/ml. Growth inhibition was inversely proportional to the concentration of the drug. DNA was extracted from cells grown in medium that contained [14C]5FU, but no carrier. The [14C]DNA was enzymatically hydrolyzed to deoxyribonucleotides, which were separated and fractionated by high-performance liquid chromatography (HPLC). The isotope was located in 2-deoxycytidine 5-monophosphate (dCMP) and 2-deoxythymidine 5-monophosphate (dTMP), with dCMP containing the majority. There was no radioactivity at the elution times for 5-fluoro-2-deoxyuridine 5-monophosphate or 2-deoxyuridine 5-monophosphate. These results suggested that 5FU was dehalogenated and the uracil moiety ultimately converted into cytosine and thymine deoxyribonucleotides. Cells were grown in [3H]uracil, and [3H]DNA was extracted and analyzed by HPLC. The isotope was found only in the pyrimidine deoxyribonucleotides, with dCMP containing 4.1 times that in dTMP. Thus, it was demonstrated that uracil and dehalogenated 5FU were not directly incorporated into DNA, but rather converted to cytosine and thymine and then incorporated into DNA by a salvage pathway.  相似文献   

2.
A quantitative analysis of JPH 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 3JPH3, 3JPH5 and 3JPH5 scalar couplings can be obtained by monitoring the intensity decay of the Pi-H3i – 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 3JPH5 and 3JPH5 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 3JPH3 couplings and relatively small 3JPH5 / 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 (DPH) as well. We will present qualitative results for the measurement of P-Hbase and P-H2 dipolar couplings in the HIV-2 TAR RNA and will discuss the reasons that so far precluded the quantification of the DPHs for the 30mer RNA.  相似文献   

3.
Random coil proton chemical shifts of deoxyribonucleic acids   总被引:2,自引:0,他引:2  
Sixteen 17-nucleotide DNA sequences have been used to determine the sequence effect on random coil DNA proton chemical shifts. Based on the proton chemical shifts measured for the central nucleotides in 64 triplets and the correction factors determined for the next nearest neighbor effects, a parameter set has been derived for predicting random coil DNA proton chemical shifts. The root-mean-square deviation (RMSD) between the predicted and the observed aromatic H6/H8 proton chemical shifts of 200 data from 22 random coil DNA sequences was determined to be 0.02 ppm with a correlation coefficient of 0.998. For the H1, H2, H2 and H3 sugar protons, the RMSD values between the predicted and the experimental shifts were found to be 0.02, 0.03, 0.03 and 0.02 ppm, respectively.  相似文献   

4.
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.  相似文献   

5.
The cohesive termini of the DNA genome of the lactococcal bacteriophage c2 were directly sequenced and appeared to be complementary, non-symmetrical, 9-nucleotide single-stranded, 3 extended DNAs, with the following sequence: 5-GTTAGGCTT-3 3-CAATCCGAA-5. DNA located on either side of the cohesive ends was sequenced and several repeats and a region with the potential for a DNA bend were found. Previously sequenced cos regions of 13 other bacteriophages were also examined for similar sequence features. All of the bacteriophages from gram-positive hosts had 3 extended DNA termini, in contrast to the bacteriophages from gram-negative hosts, which had 5 extended DNA termini. All bacteriophages had a region of dyad symmetry close to the cohesive termini. A 7.3 kb DNA fragment of the c2 genome containing the cos sequences was cloned; transduction experiments demonstrated that these cloned sequences could act as a substrate for packaging enzymes of phage c2.  相似文献   

6.
The reliability of 1H chemical shift calculations for DNA is assessed by comparing the experimentally and calculated chemical shifts of a reasonably large number of independently determined DNA structures. The calculated chemical shifts are based on semiempirical relations derived by Giessner-Prettre and Pullman [(1987) Q. Rev. Biophys., 20, 113–172]. The standard deviation between calculated and observed chemical shifts is found to be quite small, i.e. 0.17 ppm. This high accuracy, which is achieved without parameter adjustment, makes it possible to analyze the structural dependencies of chemical shifts in a reliable fashion. The conformation-dependent 1H chemical shift is mainly determined by the ring current effect and the local magnetic anisotropy, while the third possible effect, that of the electric field, is surprisingly small. It was further found that for a double helical environment, the chemical shift of the sugar protons, H2 to H5, is mainly affected by the ring current and magnetic anisotropy of their own base. Consequently, the chemical shift of these sugar protons is determined by two factors, namely the type of base to which the sugar ring is attached, C, T, A, or G, and secondly by the -angle. In particular, the H2 shift varies strongly with the -angle, and strong upfield H2 shifts directly indicate that the -angle is in the syn domain. The H1 shift is not only strongly affected by its own base, but also by its 3-neighboring base. On the other hand, base protons, in particular H5 of cytosine and methyl protons of thymine, are affected mainly by the 5-neighboring bases, although some effect (0.2 ppm) stems from the 3-neighboring base. The H2 protons are mainly affected by the 3-neighboring base. As a result of these findings a simple scheme is proposed for sequential assignment of resonances from B-helices based on chemical shifts.  相似文献   

7.
Summary 3(2)-O-glycyl-adenosine-5-monophosphate is an intermediate in the conversion of N-[imidazolyl-(1)-carbonyl]-glycine to diketopiperazine in the presence of adenosine-5-monophosphate. The significance of these observations to prebiotic chemistry is discussed.Abbreviations AMP adenosine-5-monophosphate - A adenosine  相似文献   

8.
Summary The circular DNA decamer 5-dpCGC-TT-GCG-TT-3 was studied in solution by means of NMR spectroscopy and molecular dynamics in H2O. At a temperature of 269 K, a 50/50 mixture of two dumbbell structures (denoted L2L2 and L2L4) is present. The L2L2 form contains three Watson-Crick C-G base pairs and two two-residue loops in opposite parts of the molecule. On raising the temperature from 269 K to 314 K, the L2L4 conformer becomes increasingly dominant (95% at 314 K). This conformer has a partially disrupted G(anti)-C(syn) closing base pair in the 5-GTTC-3 loop with only one remaining (solvent-accessible) hydrogen bond between NH of the cytosine dC(1) and O6 of the guanine dG(8). The opposite 5-CTTG-3 loop remains stable. The two conformers occur in slow equilibrium (rate constant 2–20 s–1). Structure determination of the L2L2 and L2L4 forms was performed with the aid of a full relaxation matrix approach (IRMA) in combination with restrained MD. Torsional information was obtained from coupling constants. Coupling constant analysis (3JHH, 3JHP, 3JCP) gave detailed information about the local geometry around backbone torsion angles , , and , revealing a relatively high flexibility of the 5-GTTC-3 loop. The values of the coupling constants are virtually temperature-independent. Weakly constrained molecular dynamics in solvent was used to sample the conformational space of the dumbbell. The relaxation matrices from the MD simulation were averaged over r–3 to predict dynamic NOE volumes. In order to account for the 1:1 conformational mixture of L2L2 and L2L4 present at 271 K, we also included S2 factors and r–6 averaging of the r–3-averaged relaxation matrices. On matrix averaging, the agreement of NOE volumes with experiment improved significantly for protons located in the thermodynamically less stable 5-GTTC-3 loop. The difference in stability of the 5-CTTG-3 and 5-GTTC-3 loops is mainly caused by differences in the number of potential hydrogen bonds in the minor groove and differences in stacking overlap of the base pairs closing the minihairpin loops. The syn conformation for dC(1), favored at high temperature, is stabilized by solvation in the major groove. However, the conformational properties of the dC(1) base, as deduced from R-factor analysis and MD simulations, include a large flexibility about torsion angle .  相似文献   

9.
Summary The carotenoid pigments of the myxobacterium Sorangium compositum were analyzed by chromatographical and chemical techniques and by visible, infra red, and mass spectroscopy. Besides -carotene, neurosporene, torulene, lycopene, and 1,2-dihydro-1-hydroxy--carotene, four new carotenoid glycosides were found. These pigments were identified as 1,2-dihydro-1-hydroxy-torulene glucoside ester (I), 1,2-dihydro-3,1-dihydroxy-torulene glucoside ester (III), 1,2-dihydro-1-hydroxy-torulene rhamnoside (II), and 1,2-dihydro-3,1-dihydroxytorulene rhamnoside (IV).Fifth communication on the carotenoids of myxobacteria. Fourth communication see Arch. Mikrobiol. 76, 364–380 (1971).  相似文献   

10.
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.  相似文献   

11.
The quality of nucleic acid solution structures can be significantly improved using residual dipolar coupling data. However, many of the one-bond couplings that could be used for this purpose are difficult to measure. Conventional 2D experiments are often unable to reveal one-bond H2-C2 and H3-C3 couplings in large RNA molecules due to spectral overlap. Here we show how to use 3D HCcH-COSY and Relay HCcH-COSY to measure one-bond H2-C2 and H3-C3 couplings which improved the precision of the structures obtained recently for a 42 nucleotide RNA.  相似文献   

12.
A model is proposed for the effect of gramicidin A on the order and structure of phospholipid dispersions. According to this model, the addition of gramicidin A influences the surrounding lipids via two independent mechanisms. The first arises from a drop in surface pressure for those lipids substantially bounded by gramicidin A. The second mechanism arises from the increase in the phospholipid headgroup spacing due to the small polar region of the polypeptide. The model provides an explanation for the currently available NMR, X-ray diffraction and Langmuir monolayer results. The model also suggests mechanisms for the ability of gramicidin A to trigger a transition of the lipid from the lamellar to hexagonal II phase, the dependence of this transition on the lipid chain length and the formation of a lamellar phase with lysophosphatidylcholine.Abbreviations NMR nuclear magnetic resonance - DMPC dimyristoylphosphatidylcholine - S molecular order parameter - CSA chemical shift anisotropy - DPPC dipalmitoylphosphati-dylcholine - LPC lysophosphatidylcholine  相似文献   

13.
Adventitious shoot regeneration was compared among leaf, stem and petal explants of carnation (Dianthus caryophyllus L.) cv. Scania on MS medium containing different concentrations of 6-benzyladenine (BA) and -naphthaleneacetic acid (NAA). High frequency regeneration was obtained only from petal explants on the media containing 5 to 10 M BA with or without 5 M NAA. Among the cytokinins tested, N-2-chloro-4-pyridyl-N-phenylurea and N-1,2,3-thiadiazol-5-yl-N-N-phenylurea were more effective than BA, kinetin, N6-2-isopentenyl adenine and zeatin on regeneration from petal explants. Although, high frequency shoot regeneration was obtained from all petal explants harvested from various developmental stages of buds, a significant decrease in regeneration capacity was observed in the explants obtained from fully-opened flowers. High frequency shoot regeneration was also obtained from the petal explants of cvs. Coral. Lena, Nora and White Sim, and an interspecific cultivar Eolo using the method developed in this study.Abbreviations NAA -naphthaleneacetic acid - BA 6-benzyladenine - GA3 gibberellic acid - 2iP N6-2-isopentenyl adenine - KT-30 N-2-chloro-4-pyridyl-N-phenylurea (also called 4PU) - TDZ N-1,2,3-thiadiazol-5-yl-N-phenylurea (also called thidiazuron)  相似文献   

14.
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 - UN 3 2-azido-2-deoxyuridine - UNH 2 2-amino-2-deoxyuridine - ImpA adenosine 5-phosphorimidazolide - ImpU uridine 5-phosphorimidazolide - ImpUN 3 2-azido-2-deoxyuridine 5-phosphorimidazolide - ImpUNH 2 2-amino-2-deoxyuridine 5-phosphorimidazolide - pA adenosine 5-phosphate - pU uridine 5-phosphate - pUN 3 2-azido-2-deoxyuridine 5-phosphate - pUNH 2 2-amino-2-deoxyuridine 5-phosphate - UpA uridylyl-[35]-adenosine - UpU uridylyl-[35]-uridine - UNpA adenylyl-[52]-2-amino-2-deoxy-uridine - UNpU uridylyl-[52]-2-amino-2-deoxyuridine (pUN)n n=2,3,4 [25]-linked oligomers of pUNH 2 poly(A) polyadenylic acid - Im imidazole - MeIm l-methylimidazole  相似文献   

15.
Crude extracts or supernatants of broken cells of Clostridium formicoaceticum reduce unbranched, branched, saturated and unsaturated carboxylates at the expense of carbon monoxide to the corresponding alcohols. The presence of viologens with redox potentials varying from E 0=-295 to-650 mV decreased the rate of propionate reduction. The more the propionate reduction was diminished the more formate was formed from carbon monoxide. The lowest propionate reduction and highest formate formation was observed with methylviologen. The carbon-carbon double bond of E-2-methyl-butenoate was only hydrogenated when a viologen was present. Formate as electron donor led only in the presence of viologens to the formation of propanol from propionate. The reduction of propionate at the expense of a reduced viologen can be followed in cuvettes. With respect to propionate Michaelis Menten behavior was observed. Experiments are described which lead to the assumption that the carboxylates are reduced in a non-activated form. That would be new type of biological reduction.Non-standard abbreviations glc Gas liquid chromatography - HPLC high performance liquid chromatography - RP reverse phase; Mediators (the figures in parenthesis of the mediators are redox potentials E 0 in mV) - CAV2+ carbamoylmethylviologen, 1,1-carbamoyl-4,4-dipyridinium dication (E 0=-296 mV) - BV2+ benzylviologen, 1,1-dibenzyl-4,4-dipyridinium dication (E 0=-360 mV) - MV methylviologen, 1,1-dimethyl-4,4-dipyridinium-dication (E 0=-444 mV) - DMDQ2+ dimethyldiquat, 4,4-dimethyl-2,2-dipyridino-1,1-ethylendication (E 0=-514 mV) - TMV2+ tetramethylviologen, 1,1,4,4-tetramethyl-4,4-dipyridinium dication (E 0=-550 mV) - PDQ2+ propyldiquat, 2,2-dipyridino-1,1-propenyl dication (E 0=-550 mV) - DMPDQ2+ dimethylpropyldiquat, 4,4-dimethyl-2,2-dipyridino-1,1-propenyl dication (E 0=-656 mV) - PN productivity number=mmol product (obtained by the uptake of one pair of electrons) x (biocatalyst (dry weight) kg)-1×h-1  相似文献   

16.
Arabidopsis consensus intron sequences   总被引:7,自引:0,他引:7  
We have analysed 998 Arabidopsis intron sequences in the EMBL database. All Arabidopsis introns to adhere to the :GU...AG: rule with the exception of 1% of introns with :GC at their 5 ends. Virtually all of the introns contained a putative branchpoint sequence (YUNAN) 18 to 60 nt upstream of the 3 splice site. Although a polypyrimidine tract was much less apparent than in vertebrate introns, the most common nucleotide in the region upstream of the 3 splice site was uridine. Consensus sequences for 5 and 3 splice sites and branchpoint sequences for Arabidopsis introns are presented.  相似文献   

17.
Purified myelin from fresh calf brain white matter was subfractionated in a discontinuous sucrose gradient; significant recovery of protein and 2,3-cyclic nucleotide 3-phosphohydrolase (CNP) and 5-nucleotidase (5N) activities occurred in all three obtained subfractions, the highest recovery being in the light subfraction; highest 5N and CNP specific activities were in medium myelin. Purified myelin was also subfractionated in a continuous sucrose gradient, with a similar localization of protein; CNP activity and 5N activity maxima suggest that myelin may be a predominant locus of 5N in bovine brain white matter. Freezing of brain white matter caused an increase in protein and in CNP and 5N total activity recoveries in denser myelin subfractions. Cytochemistry showed the reaction product of 5N in the whole myelin fraction to be associated with the innermost, outermost and medial compact myelin layers. Effects of non-ionic detergent (Lubrol WX) on 5N activity were studied, and the results also suggest the intrinsic nature of 5N as an ectoenzyme in myelin membranes. Lubrol WX was viewed as an advisable detergent for the stimulation of myelin 5N activity, but not for the solubilization of this enzyme.  相似文献   

18.
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  相似文献   

19.
Fang-Sheng Wu 《Planta》1987,171(3):346-357
The positively-charged fluorescent dye rhodamine 123 (r-123) specifically stains mitochondria in living plant protoplasts, suspensionculture cells, and root hairs. This dye functions as a vital stain and permits visualization of the localization, distribution and movement of the mitochondria. Dehydration of root hairs caused mitochondria to aggregate into clumps. Mitochondria were either homogenous or heterogeneous and were frequently seen to accumulate in the perinuclear regions of suspension-culture cells but not in those of protoplasts or root-hair cells. Dinitrophenol and high concentrations of ethyleneglycol-bis(-aminoethyl ether)-N,N,N,N-tetraacetic acid and KCl immediately eliminated fluorescence in r-123-stained mitochondria whereas ionomycin enhanced it. Treatment of seedlings with r-123 resulted in differential brightness of fluorescence in different tissues. Meristematic tissues, such as root and shoot tips, exhibited the brightest fluorescence. The cytotoxicity of r-123 in both germinating seedlings and suspension-culture cells was low. The specificity, sensitivity and low toxicity of r-123 should make it a useful tool in experiments designed to examine agents and conditions which affect the location, the physiological status or the viability of mitochondria.Abbreviations EGTA ethyleneglycol-bis-(-aminoethyl ether)N,N,N,N-tetraacetic acid - DAPI 46-diamidino-2-phenylindole - r-123 rhodamine 123  相似文献   

20.
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(Rp-S)5dT (P-thio)uridylylthymidine with theR configuration at phosphorous, and a 2,5 internucleotide linkage  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号