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1.
Yoshihiko Nishizawa 《Bioscience, biotechnology, and biochemistry》2013,77(3):229-234
During the course of an investigation of organophosphorus insecticides, the reactions of trialkyl phosphites with ethyl or phenyl trichloroacetate, diethyl monochloro- or dichloromalonate and ethyl chlorocyanoacetate were attempted. It was cleared that the above reaction did not belong to the Arbuzov reaction but to the Perkow reaction and the products had the phosphorate forms and the carbonyl group of the carboxylic acid linked with phosphorus atom by the enol-form. 相似文献
2.
Siniakov AN Butorin AS Novopashina DS Halby L Riabinin VA 《Bioorganicheskaia khimiia》2003,29(1):100-102
A new method for functionalization of oligonucleotides by addition of aminoalkyl derivatives to the intermolecular phosphorus atom of the oligonucleotide N3'-P5' phosphoramidate bond in the presence of triphenylphosphine, 4-dimethylaminopyridine, and 2,2'-dipyridyl disulfide was suggested. The reaction proceeded with both low-molecular alkylamines (1,6-diaminohexane or N,N-dimethyl-1,3-diaminopropane) and a ligand in minor groove containing a aminoalkyl group. 相似文献
3.
The phospho-intermediate formed in the reaction of the nucleoside phosphotransferase is an acyl phosphate, the phosphorus bound to the gamma-carboxylate group of a glutamic acid. Reduction of this intermediate with sodium cyanoborotritiide yields labeled 2-amino-5-hydroxyvaleric acid after hydrolysis of the protein. Nucleophilic trapping of the intermediate with hydroxylamine during the reaction with substrates leads to N-phosphohydroxylamine, which is the only reaction product at a higher concentration of hydroxylamine. Evidence is obtained from modification experiments that in addition to the carboxylate group a histidine is involved in the reaction. The pKa-value for the histidine derived from the photoinactivation of the enzyme is 7.6, indicating that this group forms a salt bridge to a carboxylate group, probably that group attacking the phosphorus. The acceptor nucleosides are bound only by hydrophobic interactions of the base, a conclusion obtained from fluorescence studies and quenching experiments. The hydrophobic interaction obviously does not involve pi-interactions to tyrosine and tryptophan residues, since their fluorescence is not affected by addition of nucleotide inhibitors. Modification of these residues leads only to unspecific inactivation. From the Scatchard plot of the titration of the enzyme with 1,N6-ethenoadenosine 5'-phosphoramidate, an efficient inhibitor (Kd = 1.2 X 10(-5) M), it can be concluded that there is only one binding site on the dimeric enzyme. 相似文献
4.
The crystal structure of the apo form of vanadium chloroperoxidase from Curvularia inaequalis reacted with para-nitrophenylphosphate was determined at a resolution of 1.5 A. The aim of this study was to solve structural details of the dephosphorylation reaction catalyzed by this enzyme. Since the chloroperoxidase is functionally and evolutionary related to several acid phosphatases including human glucose-6-phosphatase and a group of membrane-bound lipid phosphatases, the structure sheds light on the details of the dephosphorylation catalyzed by these enzymes as well. The trapped intermediate found is bound to the active site as a metaphosphate anion PO3-, with its phosphorus atom covalently attached to the Nepsilon2 atom of His496. An apical water molecule is within hydrogen-bonding distance to the phosphorus atom of the metaphosphate, and it is in a perfect position for a nucleophilic attack on the metaphosphate-histidine intermediate to form the inorganic phosphate. This is, to our knowledge, the first structural characterization of a real reaction intermediate of the inorganic phosphate group release in a dephosphorylation reaction. 相似文献
5.
Rabbit muscle aldolase in situ appears to undergo several modification reactions. One of these, specific deamidation of an asparagine residue near the COOH-terminus, appears to account for the presence of two types of subunits in the enzyme isolated from the muscle of adult rabbits. Evidence for a second modification is the presence of approximately one equivalent of organic phosphorus in the crystalline enzyme preparations. The presence of this phosphate group may be related to the incomplete release of COOH-terminal tyrosine residues from the enzyme protein with carboxypeptidase. Two reactions with substrate, both leading to the incorporation of organic phosphorus, have been demonstrated in vitro. A reaction with glyceraldehyde 3-phosphate or erythrose 4-phosphate leads to loss of catalytic activity and change in the susceptibility of COOH-terminus to carboxypeptidase. The other reaction, with fructose 1,6-diphosphate at low concentration, does not affect the activity of the enzyme, nor its susceptibility towards the action of carboxypeptidase. Either or both of these may be related to the changes which appear to occur during the life of the enzyme in vivo. 相似文献
6.
Tosisuke Hiraoka 《Histochemistry and cell biology》1973,35(4):283-296
Summary Electrophoretic means of separation revealed the presence of as many as five reaction products in Schiff-apurinic acid reaction at the maximum. They differed not only in their absorption maxima, but also in their ratios of apurinic acid phosphorus to fuchsin moiety. Some considerations on the reaction mechanism to account for the occurrence of these multiple reaction products have been made. The stoichiometry of Schiff-apurinic acid reaction was studied with respect to the main product responsible for the presentation of reaction color. A reaction product consisting of six or eight atom moles of apurinic acid phosphorus per mole of fuchsin moiety is to be formed, provided that the reagent of infinite concentration is used. From theoretical view point, a reaction product consisting of four atom moles of apurinic acid phosphorus per mole of fuchsin moiety is to be expected with the reagent of infinite concentration, provided that apurinic acid retains essentially the nucleotide sequence of its parent desoxyribonucleic acid except for some modification of the original purin nucleotide groups to react as aldehyde moieties, and provided that the reaction proceeds at a constant rate irrespective of the concentrations of the reagent. 相似文献
7.
Sinyakov A. N. Boutorine A. S. Novopashina D. S. Halby L. Ryabinin V. A. 《Russian Journal of Bioorganic Chemistry》2003,29(1):88-90
A new method for functionalization of oligonucleotides by addition of aminoalkyl derivatives to the intermolecular phosphorus atom of the oligonucleotide N3"–P5" phosphoramidate bond in the presence of triphenylphosphine, 4-dimethylaminopyridine, and 2,2"-dipyridyl disulfide was suggested. The reaction proceeded with both low-molecular alkylamines (1,6-diaminohexane or N,N-dimethyl-1,3-diaminopropane) and a minor groove binder containing an aminoalkyl group. 相似文献
8.
Gentamicin nucleotidyltransferase-catalyzed reaction of (Sp)-[alpha-17O]dATP with tobramycin produced 2"-(2'-deoxyadenosine 5'-[17O]phosphoryl)tobramycin. The configuration at phosphorus in this product was shown to be Rp by chemical degradation to chiral [17O, 18O]dAMP using a stereochemically defined procedure, and determination of the configuration at phosphorus in this product. Periodate-base treatment of 2"-(2'-deoxyadenosine 5'-[17O]phosphoryl)tobramycin followed by NaBH4 reduction produced (2-glyceryl)-[17O]dAMP, which upon snake venom phosphodiesterase-catalyzed hydrolysis in H(2)18O produced [17O,18O] dAMP. The configuration at phosphorus in this product was shown to be S by enzymatic phosphorylation to [17O,18O]dATP, adenylylcyclase (Bordetella pertussis)-catalyzed cyclization to 3',5'-cyclic [17O,18O]dAMP, and 31P NMR analysis of the ethyl esters. Since snake venom phosphodiesterase-catalyzed hydrolyses proceed with retention of configuration at phosphorus, (Sp)-[17O,18O]dAMP must have been produced from (Rp)-(2-glyceryl)-[17O]dAMP; and since the chemical degradation to the latter compound did not involve cleavage of any bonds to phosphorus, the initial enzymatic product must have been (Rp)-2"-(2'-deoxyadenosine 5'-[17O]phosphoryl)tobramycin. Therefore, nucleotidyl transfer catalyzed by gentamicin nucleotidyl-transferase proceeds with inversion of configuration at phosphorus, and the reaction mechanism involves an uneven number of phosphotransfer steps. Inasmuch as this is an uncomplicated two-substrate group transfer reaction, the mechanism probably involves direct nucleotidyl transfer from the nucleoside triphosphate to the aminoglycoside. The B. pertussis adenylylcyclase reaction was shown to proceed with inversion at phosphorus, as has been established for other adenylylcyclases. 相似文献
9.
The X-ray microprobe was examined as a possible aid in the detection of salmonellae. Results obtained indicate that specific bacterial antigen (Salmonella) increased in phosphorus and sulfur after reaction with the fluorescein isothiocyanate tagged specific antibody. Nonspecific antigen (Escherichia coli) did not increase in phosphorus and sulfur after reaction with fluorescein isothiocyanate tagged anti-Salmonella antibody. The technique developed supports the hypothesis that X-ray microprobe analysis may be useful in detecting salmonellae, or other bacteria, by determining increases in the elemental constituents of bacterial cells when reacted with elemental-tagged antibodies. 相似文献
10.
The research shows some biochemical indices in ontogenesis in broiler hen skin tissues and also its reaction to inner introduction of different doses of radioactive 137Cs. Namely, the irradiation ionizing irradiation influences actively to the indices of nuclein changes with 1 day and 1 week aged chickens. The quantity of phosphorus of nuclein acids is less with the chickens of the third group (500 Bk pr day) than with the second group (3000 Bk/day). The quantity of soluble protein, activity of the estimated fragments, the quantity of inorganic phosphorus, the concentration of ions of magnesium with chickens of the controlled group is larger than in the studied ones. It proves the influence of radionucleid 137Cs on indices which characterize the protein-nuclein change. 相似文献
11.
Almost all enzyme-catalysed phosphohydrolytic or phosphoryl transfer reactions proceed through a five-coordinated phosphorus transition state. This is also true for the phospholipase D superfamily of enzymes, where the active site usually is made up of two identical sequence repeats of an HKD motif, positioned around an approximate 2-fold axis, where the histidine and lysine residues are essential for catalysis. An almost complete reaction pathway has been elucidated by a series of experiments where crystals of phospholipase D from Streptomyces sp. strain PMF (PLD(PMF)) were soaked for different times with (i) a soluble poor, short-chained phospholipid substrate and (ii) with a product. The various crystal structures were determined to a resolution of 1.35-1.75 A for the different time-steps. Both substrate and product-structures were determined in order to identify the different reaction states and to examine if the reaction actually terminated on formation of phosphatidic acid (the true product of phospholipase D action) or could proceed even further. The results presented support the theory that the phospholipase D superfamily shares a common reaction mechanism, although different family members have very different substrate preferences and perform different catalytic reactions. Results also show that the reaction proceeds via a phosphohistidine intermediate and provide unambiguous identification of a catalytic water molecule, ideally positioned for apical attack on the phosphorus and consistent with an associative in-line phosphoryl transfer reaction. In one of the experiments an apparent five-coordinate phosphorus transition state is observed. 相似文献
12.
Alkaline phosphatase (from chicken intestinal sources) was shown to contain a considerable amount of polyanionic phosphorus which was released by basic digestion. The polyanionic phosphorus of alkaline phosphatase is not associated with protein or polyalcohols and does not exhibit a visible or ultraviolet absorption spectrum. Alkaline phosphatase and abiogenic inorganic polyphosphate were found to incorporate 32P-orthophosphate under similar experimental conditions. It has been previously reported that this enzyme will incorporate 32P-orthophosphate into its protein phosphoserine without the apparent concomitant utilization of an energy source. This reported phosphorylation was immediately reversible upon dilution of the phosphorylated enzyme with unlabelled orthophosphate, which indicates that the initial phosphorylation was an exchange reaction. These observations suggest that this polyanionic phosphorus from alkaline phosphatase may be inorganic polyphosphate. 相似文献
13.
The restriction endonuclease EcoRI hydrolyzes the Rp diastereomer of d(pGGsAATTCC), an analogue of d(pGGAATTCC) containing a chiral phosphorothioate group at the cleavage site between the deoxyguanosine and the deoxyadenosine residues (Connolly, B.A., Potter, B.V.L., Eckstein, F., Pingoud, A., and Grotjahn, L. (1984) Biochemistry 23, 3343-3453). Performing the reaction in H2(18)O leads to d(pGG) and the hexanucleotide d([18O, S]pAATTCC) which has an 18O-containing phosphorothioate group at the 5' terminus. Further hydrolysis of this hexamer with nuclease P1 yields deoxyadenosine 5'-O-[18O]phosphorothioate which can be stereospecifically phosphorylated with adenylate kinase and pyruvate kinase to give Sp-[18O] deoxyadenosine 5'-O-(1-thiotriphosphate). 31P NMR spectroscopy shows the oxygen-18 in this compound to be in a bridging position between the alpha- and beta-phosphorus atoms. Thus, the hydrolysis reaction catalyzed by EcoRI proceeds with inversion of configuration at phosphorus. This result is compatible with a direct enzyme-catalyzed nucleophilic attack of H2O at phosphorus without involvement of a covalent enzyme intermediate. 相似文献
14.
Phosphodiesterase from calf spleen exhibits nucleotidyltransferase activity when incubated with either the (PR) or the (PS) diastereomer of thymidyl 3'-(4-nitrophenyl phosphorothioate). Thymidylyl(3'-5')thymidyl phosphorothioate 3'-(4-nitrophenyl phosphorothioate) was identified as the main product of the enzyme-catalyzed reaction and the absolute configuration at the internucleotide phosphorus atom of the product was determined. The nucleotidyltransferase reaction is shown to proceed with retention of configuration at phosphorus, implying involvement of a double displacement mechanism with the formation of a nucleotidylated enzyme intermediate. To study the substrate specificity of spleen phosphodiesterase a series of deoxyribonucleosidyl 3'-(4-nitrophenyl phosphates) and phosphorothioates were synthesized, and Km and V parameters for each substrate were measured. The results obtained show virtually no specificity for substrates with different nucleosidyl moieties, while about a 20 - 30-fold drop in V and a slight increase in Km values is observed for phosphorothioate analogues as compared with corresponding phosphates. The enzyme showed no significant stereoselectivity towards phosphorothioates of opposite configurations at phosphorus. 相似文献
15.
Zhou Ning Hu Li Ming Zhou Gui Ji Liu Yan Lu Kui Zhao Yu Fen 《International journal of peptide research and therapeutics》2003,10(1):11-14
Summary The reactions of natural non-polar amino acids with phosphorus oxychloride were investigated. Some non-polar amino acids,
such as L-Phe, L-Leu and L-Val were treated with phosphorus oxychloride, then quenched with water or some bioactive compounds
(L-menthol, cinchonidine and benzylamine), and yielded the corresponding peptide and peptide conjugate libraries. ESI-MS/MS
was used to study the products of the reaction and confirm the structure of the library. This paper reports a simple method
to synthesize the homo-oligo-peptide libraries and peptide conjugated libraries. 相似文献
16.
Mevalonate-5-diphosphate decarboxylase: stereochemical course of ATP-dependent phosphorylation of mevalonate 5-diphosphate 总被引:2,自引:0,他引:2
Chicken liver mevalonate-5-diphosphate decarboxylase catalyzes the reaction of mevalonate 5-diphosphate (MVADP) with ATP to produce isopentenyl diphosphate, ADP, CO2, and inorganic phosphate. The overall reaction involves an anti elimination of the tertiary hydroxyl and carboxyl groups. To investigate the mechanism for transfer of the terminal phosphoryl group of ATP to the C-3 oxygen of MVADP, we have carried out the reaction using stereospecifically labeled (Sp)-adenosine 5'-O-(3-thio[3-17O2,18O]triphosphate) [( gamma-17O2,18O]ATP gamma S) in place of ATP. The configuration of the [17O,18O]thiophosphate produced was found to be Rp, corresponding to overall inversion of configuration at phosphorus in the thiophosphoryl group transfer step. This result is consistent with the direct transfer of the thiophosphoryl group from (Sp)-[gamma-17O2,18O]ATP gamma S to MVADP at the active site. Our result does not indicate the involvement of a covalent thiophosphoryl-enzyme on the reaction pathway. 相似文献
17.
普通小球藻生长与武汉东湖水体磷形态的相关研究 总被引:15,自引:2,他引:15
本文探讨了武汉东湖水体中总溶解磷(TSP)、溶解反应磷(SRP)、总反应磷(TRP)、溶解水解磷(SHP)和颗粒磷(PP)对普通小球藻生长的生物有效利用性。进行了普通小球藻生长量与各磷形态及总磷(TP)被利用浓度的一元相关分析,进一步讨论了这些一元相关方程的不足,并用多元回归分析建立了普通小球藻生长量(N)与所测几种磷形态被利用浓度(CTSP,CSRP,CTRP,CSHP,CPP)的相关模型。这些模型可以用于评价湖泊水体富营养化的程度及预测藻的生长潜力。 相似文献
18.
试验旨在研究低磷饲料中添加α-酮戊二酸(α-ketoglutarate, AKG)对松浦镜鲤(Cyprinus carpio Songpu)生长性能、体成分和血清生化指标的影响。选取平均体重为(6.67±0.14) g的松浦镜鲤630尾, 随机分成7组, 每组3个重复。分别投喂有效磷0.73%的正常磷饲料和有效磷0.47%的低磷饲料, 在低磷饲料中分别添加0、0.4%、0.8%、1.2%、1.6%、2.0% (干物质含量)的AKG, 配制成等氮等能饲料, 试验期为8周。结果表明: 低磷组饲料系数(FCR)显著高于正常磷组(P<0.05), 但增重率(WGR)、特定生长率(SGR)、蛋白质效率(PER)、脏体指数(VSI)及肥满度(CF)差异不显著(P>0.05); 与低磷组比, 0.4%AKG组FCR显著降低(P<0.05), 1.6%AKG组血清磷、白蛋白含量(ALB)显著增加(P<0.05); 0.8%AKG组谷草转氨酶(AST)活性和1.20%AKG组谷丙转氨酶(ALT)活性显著高于正常磷组(P<0.05); 添加不同水平AKG对血清其他指标无显著影响(P>0.05)。与正常磷组相比, 低磷组粗灰分含量、骨钙含量和钙磷沉积率显著降低(P<0.05); 与低磷组相比, 添加不同水平AKG磷沉积率显著增加(P<0.05), 0.4%AKG组, 骨钙含量显著增加(P<0.05), 0.8%AKG组, 骨磷含量显著增加(P<0.05); 低磷饲料添加AKG对其他体成分指标无显著影响(P>0.05)。由此得出, 低磷饲料中添加适量的AKG可以降低FCR, 提高松浦镜鲤脊椎骨钙磷含量。 相似文献
19.
Králíková S Masojídková M Budĕsínský M Rosenberg I 《Nucleosides, nucleotides & nucleic acids》2003,22(3):329-347
The recently described epimeric nucleosidyl-5'-C-phosphonates (alpha-hydroxyphosphonates) represent novel nucleotide analogues that can be incorporated into chimeric oligonucleotides by the phosphotriester condensation method. In order to prepare suitable protected monomer(s) we have studied condensation reaction between protected 2'-deoxythymidine and 2'-deoxythymidinyl-5'-C-phosphonate, both as model compounds, in dependence on the nature of the 5'-hydroxyl protecting group. We have found that the O-acetyl group is unstable in the presence of TPSCl or MSNT used as condensing agents for activation of the phosphorus moiety. This instability negatively influences the scope of the condensation process. On the other hand, introduction of the O-methoxycarbonyl group gave excellent results. The O-methoxycarbonyl group does not participate in the condensation process, and its quantitative introduction into the nucleotide analo gues is accomplished using a novel acylating agent, methoxycarbonyl tetrazole. 相似文献
20.
The conformation and motion of the choline head group in lipid bilayers above and below the gel-to-liquid crystal transition point are studied by means of deuterium and phosphorus magnetic resonance. For this purpose dipalmitoyl-3-sn-phosphatidylcholine is selectively deuterated at various positions on the choline and glycerol constituents. The residual deuteron quadrupole couplings and the phosphorus chemical-shift anisotropy of the corresponding lipid-water mixtures yield quantitative information on the segmental motions. The choline methyl group is only slightly hindered in its movement, but the motional freedom becomes increasingly restricted the closer the segment is located to the glycerol backbone. The average value of the OC-CN bond rotation angle changes with temperature. Increasing the temperature rotates the choline methyl group into the vicinity of the phosphorus atom. The choline group as a whole is thus characterized by a flexible, temperature-dependent structure. Its orientation in space is not fixed, either parallel or perpendicular to the bilayer surface. Instead all segments execute angular oscillations with varying degrees of restriction around the normal on the bilayer surface. The gel-to-liquid crystal phase transition at 41 degrees is clearly reflected in the deuterium and phosphorus resonance spectra of the choline moiety, while no change is observed at 34 degrees. The calorimetric pretransition at 34 degrees seems not to be associated with a conformational change in the choline group. 相似文献