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1.
Summary (3H)DIDS (4,4-diisothiocyano-2,2-ditritiostilbene-disulfonate) was used as a convalent label for membrane sites involved in anion permeability. The label binds to a small, superficially located population of sites, about 300,000 per cell, resulting in almost complete inhibition of anion exchange. The relationship of biding to inhibition is linear suggesting that binding renders each site nonfunctional. In the inhibitory range less than 1% of the label is associated with lipids but at higher concentrations of DIDS, the fraction may be as high as 4%. In ghosts, however, treatment with (3H)DIDS results in extensive labeling of lipids. In cells, a protein fraction that behavens on SDS acrylamide gels as thought its molecular weight is 95,000 daltons (95K) is predominatly labeled by (3H)DIDS. The only other labeled protein is the major sialoglycoprotein which contains less than, 5% of the total bound (3H)DIDS. Because of the linear relationship of binding to inhibition and the unique architecture of the site, it is suggested that the (3H)DIDS-binding site of the 95K protein is the substrate binding site of the anion transport system. The 95K protein is asymmetrically arranged in the membrane with the sites arranged on the outer face accessible to agent in the medium. In leaky ghost, only a few additional binding sites can be reached from the inside of the membrane in the 95K protein, in contrast to the extensive labeling of other membrane proteins in ghosts as compared to cells.Abbreviations DADS 4,4-Diamino-2,2-dihydrostilbene disulfonic acid - DIDS 4,4-Diisothiocyano-2,2-stilbene disulfonic acid - (3H)DADS 4,4-Diamino-2,2-ditritiostilbene disulfonic acid - (3H)DIDS 4,4-Diisothiocyano-2,2-ditritiostilbene disulfonic acid  相似文献   

2.
The six binary montmorillonite clay-catalyzed reactions of the5-phosphorimidazolides of adenosine, cytidine, guanosine anduridine were performed and the eight dimers from each reactionwere separated and analyzed by HPLC. A 16–51-fold higher yieldof the 5-purine-pyrimidine dimers over that of the5-pyrimidine-purines was observed. The total yield of the5-purine-pyrimidine dimers was in the 50–70% range while thatof the 5-pyrimidine-purine dimers was 1.3–7.0%. Less sequenceselectivity was observed in the homodimers formed.Regioselectivity for the formation of 3, 5-phosphodiesterbonds over that found in the absence of clay was observed. The5-purine-pyrimidine, 5-pyrimidine-pyrimidine and5-purine-purine dimers had 3, 5-links in about half of theirphosphodiester bonds. The percent phosphodiester links in the5-pyrimidine-pyrimidine dimers was 18%, a value close to thatobserved in the absence of the montmorillonite catalyst. Themontmorillonite-catalyzed reaction of all four activatednucleotides was performed and the 24 products were separated andanalyzed. The trends observed in the binary reactions wereconfirmed and the results also showed that the relativereactivity of the activated monomers was A>G>C>U in theratio 8.2: 4.8: 1.3: 1 respectively. No 5-pyrimidine-purineswith a 5-U and pG3pU, pC3pAand pC3pG weredetected. These studies suggest that a limited population ofRNAs would have formed in catalyzed prebiotic reactions.  相似文献   

3.
Summary Adenosine 5-phosphoramidates form when solutions containing adenosine 5-polyphosphates pnA (n 3) or P1, P2-diadenosine 5-diphosphate and amines are allowed to dry out. Mg ions catalyze these reactions. We have studied systems containing ammonia, imidazole, glycine, ethylenediamine and histamine. The yields of adenosine 5-phosphoramidates range from 10–50 % based on the nucleotide. The prebiotic significance of the reactions is discussed.Abbreviations Im imidazole - hist histamine - gly glycine - en ethylenediamine - CDI 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride - EDTA ethylenediaminetetraacetic acid - A adenosine - Pn (n = 1, 2 ) linear polyphosphate containing n phosphate residues - pnA adenosine 5-polyphosphate containing n phosphate residues - ADP adenosine 5-diphosphate - ATP adenosine 5-triphosphate - AppA P1, P2-diadenosine 5-diphosphate - gly-pA adenylyl-(5N)-glycine - ImpA adenosine 5-phosphorimidazolide - NH2-pA adenosine 5-phosphoramidate - en-pA adenylyl-(5N)-ethylenediamine - hist (NH) - pA adenosine 5-phospho-[2-(4-imidazolyl)-ethylamide] - hist(Im)-pA adenosine 5-phospho-[4-(2-aminoethyl)-imidazolide] - enP1,2 phosphoramidates of ethylenediamine derived from H3PO4 and H4P2O7  相似文献   

4.
Females of the medfly, Ceratitis capitata, prefer sucrose solutions containing ribonucleotides to sucrose solutions without them. The order of preference for the nucleotides was: 5GMP>GTP>5CMP>5IMP >dGMP>5UMP>5AMP>5XMP=ATP=2 & 3GMP=RP>3AMP.2AMP, guanosine, inosine, adenine and 5TMP produced no significant stimulation. Females sterilized by irradiation showed reduced attraction to 5GMP as compared to non-irradiated females.Optimal molecular configuration for phagostimulation includes: phosphorylation at the 5 position of the ribose, free hydroxyl groups at 2 and 3 on the ribose, and an NH2 group at the 2 position of the aromatic ring of purine.It is proposed that the 5GMP in yeast hydrolyzate can be used as a measure of the suitability of the hydrolyzate as a bait.
Résumé La femelle de la mouche méditerranéenne des fruits, Ceratitis capitata, préfère les solutions de sucrose contenant des ribonucléotides aux simples solutions de sucrose. Lórdre de préférence pour les nucléotides est le suivant: 5GMP>GTP>5CMP>5IMP >dGMP>5UMP>5AMP>5XMP=ATP =2 & 3GMP=RP>3AMP.Le 2AMP, la guanosine, l'inosine, l'adénine et le 5TMP provoquent une stimulation significative. Les femelles montrent aprés stérilisation par irradiation une attirance réduite pour le 5GMP par comparaison avec les femelles non-irradiées.La configuration moléculaire optimale pour la phagostimulation comprend: la phosphorylation en position 5 du ribose; des groupes hydroxyles libres en 2 et 3 sur le ribose; et un groupe NH2 en position 2 sur le noyau aromatique.Nous proposons que le 5GMP dans l'hydrolysat de levure puisse être utilisé pour mesurer la capacité de l'hydrolysat comme appât.

Abbreviations 5AMP Adenosine 5-monophosphate - 3AMP Adenosine 3-monophosphate - 2AMP Adenosine 2-monophosphate - dAMP 2-deoxyadenosine 5-monophosphate - ADP Adenosine 5-diphosphate - ATP Adenosine 5-triphosphate - 5GMP Guanosine 5-monophosphate - 2GMP Guanosine 2-monophosphate - 3GMP Guanosine 3-monophosphate - dGMP 2-deoxyguanosine 5-monophosphate - GDP Guanosine 5-diphosphate - GTP Guanosine 5-triphosphate - 5IMP Inosine 5-monophosphate - IDP Inosine 5-diphosphate - ITP Inosine 5-triphosphate - 5XMP Xanthosine 5-monophosphate - 5CMP Cytidine 5-monophosphate - dCMP 2 deoxycytidine 5-monophosphate - CTP Cytidine 5-triphosphate - 5UMP Uridine 5-monophosphate - 5TMP Thymidine 5-monophosphate - RP Ribose 5 monophosphate  相似文献   

5.
In the course of characterization of glycolipid sulfotransferase from human renal cancer cells, the manner of inhibition of sulfotransferase activity with pyridoxal 5-phosphate was investigated. Incubation of a partially purified sulfotransferase preparation with pyridoxal 5-phosphate followed by reduction with NaBH4 resulted in an irreversible inactivation of the enzyme. When adenosine 3-phosphate 5-phosphosulfate was co-incubated with pyridoxal 5-phosphate, the enzyme was protected against this inactivation. Furthermore, pyridoxal 5-phosphate was found to behave as a competitive inhibitor with respect to adenosine 3-phosphate 5-phosphosulfate with aK i value of 287 µm. These results suggest that pyridoxal 5-phosphate modified a lysine residue in the adenosine 3-phosphate 5-phosphosulfate-recognizing site of the sulfotransferase.  相似文献   

6.
The concept of self-consistent J coupling evaluation exploits redundant structure information inherent in large sets of 3J coupling constants. Application to the protein Desulfovibrio vulgaris flavodoxin demonstrates the simultaneous refinement of torsion-angle values and related Karplus coefficients. The experimental basis includes quantitative coupling constants related to the polypeptide backbone torsion originating from a variety of heteronuclear 2D and 3D NMR correlation experiments, totalling 124 3J(HN,H), 129 3J(HN,C), 121 3J(HN,C), 128 3J(Ci–1,Hi), 121 3J(Ci–1,Ci), and 122 3J(Ci–1,Ci). Without prior knowledge from either X-ray crystallography or NMR data, such as NOE distance constraints, accurate dihedral angles are specified for 122 non-glycine and non-proline residues out of a total of 147 amino acids. Different models of molecular internal mobility are considered. The Karplus coefficients obtained are applicable to the conformational analysis of torsions in other polypeptides.  相似文献   

7.
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   

8.
Summary In the human proteolipid protein gene, the base sequence of the intronic region 5 to exon 6 was found to be 5-ctctttcattttcctgcag-3 and not 5-ctctttt-cattttcctgcag-3 as previously reported.  相似文献   

9.
Summary The activity of poly (ADP-ribose) polymerase (ADPRP) and the content of 2,5-oligodenylates core (2,5An; n = 2,3 and 4) were measured in homogenates of the uterus and of the liver of immature rats immediately before (time 0) or at different times after injection of estradiol-valerate. ADPRP activity increased gradualy, starting 6 hours after estrogen injection, for about 4 days. Instead, the content of 2,5 An decreased by about 50% within 6 hours, and thereafter more slowly for 4 days to about 20% of starting values. Estrogen increased ADPRP activity and decreased 2,5An concentration also in the kidney and in the cardiac muscle of the same animals, but not in the skeletal muscle, where neither of the two parameters was affected. Injection of vehicle only (sesame oil) had no effect on ADPRP activity nor on 2,5An content of immature rat tissues.  相似文献   

10.
Membrane preparations, capable of high rates of respiration-linked ATP synthesis, have been obtained from a gram-positive methylotrophic bacterium Bacillus sp. MGA3. NADH, succinate, reduced TMPD and methanol were shown to be suitable substrates for the oxidative phosphorylation. Esterification of orthophosphate was dependent on electron transfer, as evidenced by the requirement for both substrate and oxygen. Phosphorylation was also dependent on ADP and was destroyed by boiling the membrane preparation. The phosphorylation was markedly uncoupled by carbonyl cyanide p-(trichloromethoxy)-phenylhydrazone (CCCP) and was inhibited by N,N-dicyclohexylcarbodiimide (DCCD). KCN caused strong inhibition of substrate oxidation as well as phosphorylation for all substrates tested. Rotenone, amytal and antimycin A caused inhibition when NADH or methanol were used as substrates. Antimycin A inhibited respiration and ATP synthesis with succinate as substrate and had no effect on ascorbate —N,N,N,N-tetramethyl-p-phenylenediimide (TMPD) oxidation by membrane preparations of Bacillus sp. MGA3. P/O ratios determined were 2.4 with NADH, 1.7 with succinate and 0.8 with reduced TMPD. The measured P/O ratio with methanol-oxidizing system was similar to that with NADH (about 2.4).Abbreviations CCCP Carbonyl cyanide p-(trichloromethoxy)-phenylhydrazone - DCCD N,N-dicyclohexylcarbodiimide - TMPD N,N,N,N-tetramethyl-p-phenylenediimide - Q ubiquinone Q  相似文献   

11.
The specific binding of vasoactive intestinal peptide (VIP) to bovine thyroid plasma membranes is inhibited by guanine nucleotides. Guanosine 5-triphosphate (GTP) and the non-hydrolyzable GTP analogs guanosine 5-,-imidotriphosphate (Gpp(NH)p) and guanosine 5-O-(3-thiotriphosphate) (GTP--S) inhibited markedly the binding of VIP to its receptors. This inhibition was higher with GTP than with Gpp(NH)p and GTP--S and was due to an increase of the rate of dissociation of peptide bound to membranes. Other nucleotides did not show any effect.  相似文献   

12.
The white rot basidiomycete Phanerochaete chrysosporium metabolized 4-ethoxy-3-methoxyphenyl-glycerol--guaiacyl ether (V) in low nitrogen, stationary cultures under which conditions the ligninolytic enzyme system is expressed. 4-Ethoxy-3-methoxyphenylglycerol XIII, guaicol and 4-ethoxy-3-methoxybenzyl alcohol (II) were isolated as metabolic products. Exogenously added XIII was rapidly converted to 4-ethoxy-3-methoxybenzyl alcohol indicating that it is an intermediate in the metabolism of V. P. chrysosporium also metabolized 1-(4-ethoxy-3-methoxyphenyl)-2-(2-methoxyphenoxy)-3-hydroxypropane VI. The degradation pathway for this dimer also included initial -ether cleavage and -hydroxylation of the diol product 1-(4-ethoxy-3-methoxyphenyl) 2,3 dihydroxypropane (XI) to yield the triol XIII which was cleaved at the , bond to yield 4-ethoxy-3-methoxybenzyl alcohol. Finally P. chrysosporium also cleaved the dimer 1-(4-ethoxy-3-methoxyphenyl)-2-(2-methoxyphenoxy)-1-hydroxypropane (VIII) at the -ether linkage yielding 1-(4-ethoxy-3-methoxyphenyl) 1,2 dihydroxypropane (IX) which was subsequently cleaved at the , bond to yield II. All of the results indicate that oxidative -ether cleavage is an important initial reaction in the metabolism of -aryl ether lignin substructure dimeric compounds. Metabolities were identified after comparison with chemically synthesized standards by gas liquid chromatography-mass spectrometry.Abbreviations GLC Gas liquid chromatography - TMSi trimethylsilyl - TLC thin layer chromatography  相似文献   

13.
Experimental results are presented for the verification of the specific interaction step of the adsorbed template biogeochemical cycle, a simple model for a primitive prebiotic replication system. The experimental system consisted of gypsum as the mineral to which an oligonucleotide template attaches (Poly-C or Poly-U) and 5-AMP, 5-GMP, 5-CMP and 5-UMP as the interacting biomonomers. When Poly-C or Poly-U were used as adsorbed templates, 5-GMP and 5-AMP, respectively were observed to be the most strongly adsorbed species. Moreover, there exists a direct quantitative relationship between the quantity of cytidine or uracil residues in the adsorbed state and the amount of the complementary mononucleotide that is attached to it. NaCl added to the system in order to create conditions of high ionic strength seems to enhance the selectivity of the adsorption of the monmucleotides to these adsorbed templates.  相似文献   

14.
Summary The FhuA protein in the outer membrane of Escherichia coli serves as a multifunctional receptor for the phages T5, T1, 80, for colicin M, for ferrichrome (Fe3+-siderophore) and for the structurally related antibiotic, albomycin. To determine structural domains required for these receptor functions and for export, a fusion protein between FhuA and Iut (receptor for Fe3+-aerobactin and cloacin DF13) was constructed. In the FhuA-Iut hybrid protein, 24 amino acids of FhuA were replaced by 19 amino acids, 18 of which were from Iut. The number of plaque forming units of phage T5 and T1 on cells expressing FhuA-Iut was nearly as high as on cells expressing plasmid-encoded wild-type FhuA. However, 107-fold higher concentrations of phage 80 and 103 times more colicin M were required to obtain a zone of growth inhibition. Truncated FhuA proteins in which the last 24 amino acids at the carboxy-terminus were replaced by 16 (FhuA2) or 3 (FhuAT) amino acids could hardly be detected on polyacrylamide electrophoretograms of outer membrane proteins, due to proteolytic degradation. Sensitivity of cells expressing FhuA2 to phage T5 and T1 was reduced by several orders of magnitude and sensitivity to phage 80 and colicin M was totally abolished. In contrast, cells expressing FhuAT were nearly as sensitive to phage T5, T1, and 80 and to colicin M as cells containing FhuA-Iut. None of the constructs could grow on ferrichrome as sole iron source and none was sensitive to albomycin. Ferrichrome did not inhibit binding of T5 to TonB cells expressing FhuA-Iut (as it did in FhuA+ cells) due to the lack of ferrichrome binding. It is concluded that very small deletions (relative to the size of FhuA with 714 amino acids) at the C-terminal end render FhuA susceptible to proteolytic cleavage. The C-terminal alterations affect sensitivity to FhuA-specific agents very differently. Apparently, the C-terminus is a very important part of FhuA regarding stability and activity. Expression of active FhuA and partially inactive FhuA derivatives in the same cell revealed no negative complementation, suggesting a FhuA monomer as functional unit.  相似文献   

15.
Plausible prebiotic conditions for the phosphorylation of nucleosides by inorganic phosphate were reported by Lohrmann and Orgel in 1971. This reaction was carried out on heated dry films and promoted by urea. The major products formed were nucleoside-2:3 cyclicPs; 5-NMPs and other derivatives were also formed. Minor modifications of the Lohrmann and Orgel system have resulted in the preferential formation of 5-NMPs. In this modified system a 2-fold preference for phosphorylation of the 5-OH group over the 3(2)-OH group was observed and the formation of other derivatives was minimized. The small amounts of bis compounds that were formed in this system could be quantitatively removed by selective binding to the mineral hydroxylapatite at moderate ionic strengths. It was also discovered that under hydrolytic conditions there was a 3:1 preference for removal of phosphates attached to the 3-OH group over the 5-OH group. A recycling procedure for obtaining additional 5-NMPs from bis compounds and 3-NMPs is proposed.  相似文献   

16.
Adenosine-5-triphosphate was synthesized by the phosphorylation of adenosine-5-diphosphate in aqueous solution containing cyanate as a condensing reagent and insoluble calcium phosphate produced from phosphate and calcium chloride. In a similar manner, adenosine-5-diphosphate was synthesized from adenosine-5-monophosphate. When the experiment was carried out in the conditions of 4 °C and pH 5.75, the formation of adenosine-5-diphosphate and adenosine-5-triphosphate from adenosine-5-monophosphate was observed in the yields of 19 and 7%, respectively. The other nucleoside-5-triphosphates were also produced from their respective diphosphates.  相似文献   

17.
Internal motions of d-ribose selectively 2H-labeled at the 2 position were measured using solid state 2H NMR experiments. A sample of d-ribose-2 -d was prepared in a hydrated, non-crystalline state to eliminate effects of crystal-packing. Between temperatures of –74 and –60°C the C2–H2 bond was observed to undergo two kinds of motions which were similar to those of C2–H2/H2 found previously in crystalline deoxythymidine (Hiyama et al. (1989) J. Am. Chem. Soc., 111, 8609–8613): (1) Nanosecond motion of small angular displacement with an apparent activation energy of 3.6 ± 0.7 kcal mol–1, and (2) millisecond to microsecond motion of large amplitude with an apparent activation energy 4 kcal mol–1. At –74°C, the slow, large-amplitude motion was best characterized as a two-site jump with a correlation time on the millisecond time scale, whereas at –60°C it was diffusive on the microsecond time scale. The slow, large-amplitude motions of the C2–H2 bond are most likely from interconversions between C2-endo and C3-endo by way of the O4-endo conformation, whereas the fast, small-amplitude motions are probably librations of the C2–H2 bond within the C2-endo and C3-endo potential energy minima.  相似文献   

18.
Studies on the molecular basis of H+ translocation by cytochromec oxidase   总被引:2,自引:0,他引:2  
We report here studies which characterize further the interaction ofN,N-dicyclohexylcarbodiimide with cytochromec oxidase leading to inhibition of H+ translocation by the enzyme. Further evidence is presented to show that the inhibition results from a real interaction of DCCD with the enzyme and cannot be accounted for by uncoupling and, contrary to recent criticisms, this interaction occurs specifically with subunit III of the enzyme even at relatively high inhibitor-to-enzyme stoichiometries. Use of a spin-label analogue of DCCD has enabled us to demonstrate that the carbodiimide-binding site is highly apolar and may not lie on the pathway of electron transfer.Abbreviations DCCD N,N-dicyclohexylcarbodiimide - NCCD N-(2, 2, 6, 6-tetramethylpiperidyl-1-oxyl)-N-(cyclohexyl)carbodiimide - Hepes 2-(N-2-hydroxyethylpiperazin-N-yl) ethane sulfonate - TMPD N,N,N,N-tetramethylphenylenediamine  相似文献   

19.
Summary Strain 211-1aM of Saccharomyces cerevisiae was found to specifically incorporate into its nuclear and mitochondrial DNA exogenous 5-dTMP when it is incubated in a synthetic medium N rich in inorganic phosphate. 3-digestions of DNA from cells labelled with 32P-5-dTMP revealed that the 5-dTMP molecules pass through cell walls and membranes to the sites of DNA synthesis without being broken down. It is possible in medium N to generate DNA which almost totally derives its thymine-contents from external sources when rather high concentrations of 5-dTMP (approx. 100 g/ml) are offered.  相似文献   

20.
Relationships of vegetation, climate and soils in Shanxi plateau wereanalyzed by use of Canonical Correspondence Analysis (CCA). Shanxi province,located at 34°35–40°43 N, 110°15–114°33 E, was divided into a series of rectangular districts of30 latitude by 20 longitude. Areas of vegetation formations and soil types ineach district were measured carefully using fine grain on the vegetation andsoil maps of Shanxi. Climatic data were mean values of 25 years records in eachdistrict. Three data matrices of climate, vegetation and soil were combined byCCA. The results showed that the distribution of vegetation is closely relatedto the variety of climates and to soils distribution.  相似文献   

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