84-52-6Relevant articles and documents
Isothermal titration calorimetric study of RNase-A kinetics (cCMP → 3′-CMP) involving end-product inhibition
Spencer, Shawn D.,Raffa, Robert B.
, p. 1642 - 1647 (2004)
Purpose. Isothermal titration calorimetry (ITC) and progress curve analysis was used to measure the enzyme kinetic parameters (K M and k cat) of the hydrolysis of cCMP by RNase-A, a reaction that includes end-product competitive inhibition by 3'-CMP. Methods. The heat generated from injection of 9-15 μl cCMP (20 mM) into bovine pancreatic RNase-A (600 nM) in 50 mM Na+ acetate buffer (pH 5.5; 37°C) was monitored for 1500-2000 s. Thermal power (dQ/dt), equal to (1) /ΔH app × d(cCMP)/dt was recorded every 1 s. The end-product inhibition constant (K p) and enthalpy of the inhibitor binding interaction was obtained from the saturation data of 60 sequential injections of 3′-CMP (1.2 mM) into 0.05 mM RNase-A. The data of the plot of -d[cCMP]/dt against [cCMP] were fitted to kinetic equations incorporating K p to yield K M and k cat. Results. ΔH app for each run was obtained by integration of the progress curve. The plot of -d[cCMP]/dt against [cCMP] yielded the kinetic parameters K M = 105.3 μM, 121.6 μM, and 131.3 μM; k cat = 1.63 s-1, 1.56 s-1, and 1.71 s-1. The end-product bound with 1:1 stoichiometry and K p = 53.2 μM. Conclusions. The combination of progress curve analysis and ITC allowed rapid and facile measurement of the kinetic parameters for catalytic conversion of cCMP to 3′-CMP by RNase-A, a reaction complicated by end-product inhibition.
A nearly isosteric photosensitive amide-backbone substitution allows enzyme activity switching in ribonuclease S
Wildemann, Dirk,Schiene-Fischer, Cordelia,Aumueller, Tobias,Bachmann, Annett,Kiefhaber, Thomas,Luecke, Christian,Fischer, Gunter
, p. 4910 - 4918 (2007)
ψ[CS-NH]4-RNase S, a site specific modified version of RNase S obtained by thioxylation (O/S exchange) at the Ala4-Ala 5- peptide bond, was used to evaluate the impact of protein backbone photoswitching on bioactivity. ψ[CS-NH]4-RNase S was yielded by recombination of the S-protein and the respective chemically synthesized thioxylated S-peptide derivative. Comparison with RNase S revealed similar thermodynamic stability of the complex and an unperturbed enzymatic activity toward cytidine 2′,3′-cyclic monophosphate (cCMP). Reversible photoisomerization with a highly increased cis/trans isomer ratio of the thioxopeptide bond of ψ[CS-NH]4-RNase S in the photostationary state occurred under UV irradiation conditions (254 nm). The slow thermal reisomerization (t1/2 = 180 s) permitted us to determine the enzymatic activity of cis ψ[CS-NH]4-RNase S by measurement of inital rates of cCMP hydrolysis. Despite thermodynamic stability of cis ψ[CS-NH]4-RNase S, its enzymatic activity is completely abolished but recovers after reisomerization. We conclude that the thioxopeptide bond modified polypeptide backbone represents a versatile probe for site-directed photoswitching of proteins.
Guanidine-based polymer brushes grafted onto silica nanoparticles as efficient artificial phosphodiesterases
Savelli, Claudia,Salvio, Riccardo
, p. 5856 - 5863 (2015/03/31)
Polymer brushes grafted to the surface of silica nanoparticles were fabricated by atom-transfer radical polymerization (ATRP) and investigated as catalysts in the cleavage of phosphodiesters. The surfaces of silica nanoparticles were functionalized with an ATRP initiator. Surface-initiated ATRP reactions, in varying proportions, of a methacrylate moiety functionalized with a phenylguanidine moiety and an inert hydrophilic methacrylate species afforded hybrid nanoparticles that were characterized with potentiometric titrations, thermogravimetric analysis, and SEM. The activity of the hybrid nanoparticles was tested in the transesterification of the RNA model compound 2-hydroxypropyl para-nitrophenylphosphate (HPNP) and diribonucleoside monophosphates. A high catalytic efficiency and a remarkable effective molarity, thus overcoming the effective molarities previously observed for comparable systems, indicate the existence of an effective cooperation of the guanidine/guanidinium units and a high level of preorganization in the nanostructure. The investigated system also exhibits a marked and unprecedented selectivity for the diribonucleoside sequence CpA. The results presented open up the way for a novel and straightforward strategy for the preparation of supramolecular catalysts.
COMPOUNDS FOR IMMUNOPOTENTIATION
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Page/Page column 151, (2010/02/15)
Methods of stimulating an immune response and treating patients responsive thereto with 3,4-di(1H-indol-3-yl)-1H-pyrrole-2,5-diones, staurosporine analogs, derivatized pyridazines, chromen-4-ones, indolinones, quinazolines, nucleoside analogs, and other small molecules are disclosed.
Dinuclear Zn2+ complexes in the hydrolysis of the phosphodiester linkage in a diribonucleoside monophosphate diester.
Yashiro, Morio,Kaneiwa, Hideki,Onaka, Kenichi,Komiyama, Makoto
, p. 605 - 610 (2007/10/03)
Dizinc complexes that were formed from 2:1 mixtures of Zn(NO3)2 and dinucleating ligands TPHP (1), TPmX (2) or TPpX (3) in aqueous solutions efficiently hydrolyzed diribonucleoside monophosphate diesters (NpN) under mild conditions. The dinucleating ligand affected the structure of the aquo-hydroxo-dizinc core, resulting in different characteristics in the catalytic activities towards NpN cleavage. The pH-rate profile of ApA cleavage in the presence of (Zn2+)(2)-1 was sigmoidal, whereas those of (Zn2+)(2)-2 and (Zn2+)(2)-3 were bell-shaped. The pH titration study indicated that (Zn2+)(2)-1 dissociates only one aquo proton (up to pH 12), whereas (Zn2+)(2)-2 dissociates three aquo protons (up to pH 10.7). The observed differences in the pH-rate profile are attributable to the various distributions of the monohydroxo-dizinc species, which are responsible for NpN cleavage. As compared to that using (Zn2+)(2)-1, the NpN cleavage using (Zn2+)(2)-2 showed a greater rate constant, with a higher product ratio of 3'-NMP/2'-NMP. The saturation behaviors of the rate, with regard to the concentration of NpN, were analyzed by Michaelis-Menten type kinetics. Although the binding of (Zn2+)(2)-2 to ApA was weaker than that of (Zn2+)(2)-1, (Zn2+)(2)-2 showed a greater kcat value than (Zn2+)(2)-1, resulting in higher ApA cleavage activity of the former.
The pKa of the internucleotidic 2′-hydroxyl group in diribonucleoside (3′→5′) monophosphates
Acharya,Foeldesi,Chattopadhyaya
, p. 1906 - 1910 (2007/10/03)
Ionization of the internucleotidic 2′-hydroxyl group in RNA facilitates transesterification reactions in Group I and II introns (splicing), hammerhead and hairpin ribozymes, self-cleavage in lariatRNA, and leadzymes and tRNA processing by RNase P RNA, as well as in some RNA cleavage reactions promoted by ribonucleases. Earlier, the pKa of 2′-OH in mono- and diribonucleoside (3′-5′) monophosphates had been measured under various nonuniform conditions, which make their comparison difficult. This work overcomes this limitation by measuring the pKa values for internucleotidic 2′-OH of eight different diribonucleoside (3′-5′) monophosphates under a set of uniform noninvasive conditions by 1H NMR. Thus the pKa is 12.31 (±0.02) for ApG and 12.41 (±0.04) for ApA, 12.73 (±0.04) for GpG and 12.71 (±0.08) for GpA, 12.77 (±0.03) for CpG and 12.88 (±0.02) for CpA, and 12.76 (±0.03) for UpG and 12.70 (±0.03) for UpA. By comparing the pKas of the respective 2′-OH of monomeric nucleoside 3′-ethyl phosphates with that of internucleotidic 2′-OH in corresponding diribonucleoside (3′→5′) monophosphates, it has been confirmed that the aglycons have no significant effect on the pKa values of their 2′-OH under our measurement condition, except for the internucleotidic 2′-OH of 9-adeninyl nucleotide at the 5′-end (ApA and ApG), which is more acidic by 0.3-0.4 pKα units.
Phosphodiester Cleavage of Ribonucleoside Monophosphates and Polyribonucleotides by Homo- and Heterodinuclear Metal Complexes of a Cyclohexane-Based Polyamino-Polyol Ligand
Jancso, Attila,Mikkola, Satu,Loennberg, Harri,Hegetschweiler, Kaspar,Gajda, Tamas
, p. 5404 - 5415 (2007/10/03)
The ability of the dinuclear complexes of tdci [1,3,5-trideoxy-1,3,5-tris(dimethylamino)-cis-inositol] to promote the cleavage of the phosphodiester bonds of nucleoside 2′,3′-cyclic monophosphates, dinucleoside monophosphates and polyribonucleotides has been studied. The homodinuclear copper(II) and zinc(II) complexes efficiently promote the hydrolysis of cyclic nucleotides. The second-order rate constant (k2≈0.44M-1S-1) estimated for the cleavage of 2′,3′-cAMP induced by dinuclear copper(II) complexes is about 107 times greater than that for the hydroxide-ion-catalysed reaction. The complex selectively cleaves the 2′O-P bond of 2′,3′-cUMP and forms the 3′-product in 91% yield. An equimolar mixture of copper(II), zinc(II) and tdci proved to be more efficient than either of the binary systems: a 7-20-fold rate enhancement was observed for the cleavage of 2′,3′-cNMP substrates. The half-life for the hydrolysis of 2′,3′-cAMP decreased from 300 days to five minutes at 25°C when the concentration of each of the three components was 2.5mM. In contrast to the copper(II) or zinc(II) complexes of tdci, the heterodinuclear species promoted the hydrolysis of several dinucleoside monophosphates. For two ApA isomers, cleavage of the 3′,5′-bond was about 6.5 times faster than cleavage of the 2′,5′-bond. On the basis of the kinetic data, a trifunctional mechanism is suggested for the heterodinuclear-complex-promoted cleavage of the phosphodiester bond. Double Lewis acid activation occurs when the metal ions bind to the phosphate oxygen atoms. In particular, a metal-bound hydroxide ion serves as a general base or a nucleophilic catalyst, and, presumably, a zinc(II)-bound aqua ligand behaves as a general acid and facilitates the departure of the leaving alkoxide group. The effect of the complexes on the hydrolysis of poly(U), poly(A) and type III native RNA was also investigated, and, for the first time, kinetic data on the cleavage of the phosphodiester bonds of polyribonucleotides by a dinuclear complex was obtained.
Rapid and highly base selective RNA cleavage by a dinuclear Cu(II) complex
Liu, Shanghao,Hamilton, Andrew D.
, p. 587 - 588 (2007/10/03)
A bis-Cu(II) complex based on a covalently linked terpyridine and bipyridine ligand system is shown to rapidly cleave bis-ribonucleotides with remarkable selectivity for adenine bases.
Efficient RNA hydrolysis by lanthanide(III)-hydrogen peroxide combinations. Novel aggregates as the catalytic species
Kamitani, Jun,Sumaoka, Jun,Asanuma, Hiroyuki,Komiyama, Makoto
, p. 523 - 527 (2007/10/03)
Combinations of lanthanum(III) ion and hydrogen peroxide efficiently hydrolyze RNA under physiological conditions, because of a synergetic cooperation. The rate constant for the hydrolysis of adenylyl(3′-5′)adenosine at pH 7.2 and 30°C is 7.7 x 10-2 min-1, when [LaIII]0 = 10 and [H2O2]0 = 100 mM. This value is 460 times as great as that for the ApA hydrolysis by La alone (1.7 x 10-4 min-1). Hydrogen peroxide is inactive when used separately. A similar synergism operates between NdIII and H2O2. According to the kinetic analysis and the potentiometric titration, a trimeric aggregate of [La(O-O)3La] complex is responsible for the RNA hydrolysis. This result is in contrast with the previous proposal on the hydrolysis of bis(4-nitrophenyl)phosphate that monomeric species of [La(O-O)2La]2+ is the active species (B. K. Takasaki and J. Chin, J. Am. Chem. Soc., 1995, 117, 8582). The discrepancy is ascribed to the difference in the basicities of the leaving groups in the substrates.
Rapid and highly selective cleavage of ribonucleoside 2',3'-cyclic monophosphates by dinuclear Cu(II) complexes
Liu,Luo,Hamilton
, p. 2678 - 2680 (2007/10/03)
Two characteristics that never before appeared together are combined in complexes 1 and 2, which show high activity and high selectivity in the hydrolysis of cyclic nucleoside 2',3'-monophosphates as model compounds for RNA. In the case of 1 the regioselectivity is exceptional, and, in the case of 2, the base selectivity.
