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E Schlimme

Publications and source records attributed to E Schlimme.

At least 37 records · Page 2Linked to original sources
PubMed1984

Development of a chromatographic method for the quantitative determination of minor ribonucleosides in physiological fluids. Characterization and quantitative determination of minor ribonucleosides in physiological fluids, Part I.

We describe an on-line multi-column high performance liquid chromatographic method for the selective clean-up and analysis of major and minor ribonucleosides in physiological fluids. Quantitative data obtained for the determination of some methylated ribonucleosides in human urines are compared with those obtained with the traditional off-line method. The on-line technique developed in our laboratory is distinguished from the off-line method by the following features: Sample clean-up and analysis of the target-compounds can easily be automatized, Total time of analysis, for example of urinary ribonucleosides, is decreased to 35 minutes, Laborious and error-prone evaporation and redissolution steps are avoided, Reliability of the overall analytical system can be controlled with ease, Small sample-volumes can be applied directly, Sensitive samples can be processed very rapidly under mild conditions, Results obtained with the on-line and off-line-techniques compare well.

Boronic Acids↗
PubMed1983

On-line high-performance liquid affinity chromatography-high-performance liquid chromatography analysis of monomeric ribonucleoside compounds in biological fluids.

We describe an on-line multi-dimensional column chromatography system. It consists of a high-performance liquid affinity chromatography perfractionation column for cis-diol compounds and a series-connected reversed-phase high-performance liquid chromatography system for the analysis of ribonucleosides. This on-line procedure allows the rapid and direct analysis of methylated ribonucleic acid catabolites in biological fluids (serum, urine) and therefore might be useful in pathobiochemistry.

Chemical Phenomena↗
PubMed1983

A model for metalloprotein-catalysed ADP, ATP transport in mitochondria.

We propose a hypothetical model for the transmembrane exchange reaction catalysed by the mitochondrial adenine nucleotide carrier protein, which basically consists of an alternating reorientation of a transitory carrier-metal-nucleotide complex. The key features of the model are: the participation of an intrinsic divalent metal ion in the course of transport catalysis; the different stability constants of protonated and deprotonated nucleotide-metal complexes; the exposure and retraction of strategic arginyl residues; the alternating reorientation of the active center involving a change from the cytosolic conformation (Cc) to the matrix conformation (Cm).

Animals↗
PubMed1983

Inhibition study of ADP,ATP transport in mitochondria with trinitrophenyl-modified substrates.

The ADP,ATP carrier of rat liver mitochondria is specifically inhibited by Meisenheimer-type trinitrophenyl (TNP) derivatives of ADP and ATP. Due to a systematic inhibition study we could show that the TNP-moiety itself, even in the 2', 3'-O-cyclic Meisenheimer complex, revealed no inhibition of mitochondrial ADP,ATP transport. Nucleosidic TNP-compounds are weak inhibitors. Introduction of a phosphate group at the 5'-position, however, enhances the inhibitory power markedly. Our findings point to a synergistic effect of the 5'-phosphate chain and the TNP-moiety. Irreversible inhibition by TNP-nucleotides can be ruled out to the fact that the inhibited ADP,ATP-transport system is fully reactivated by addition of albumin.

Adenosine Diphosphate↗
PubMed1980

Adenosine di- and triphosphate transport in mitochondria. Role of the amidine region for substrate binding and transport.

A variety of base-modified nucleotide analogues was prepared and characterized as their alpha-32P- or U-14C-labeled compounds. Carrier-linked nucleotide binding and carrier-catalyzed exchange across the inner membrane of rat liver mitochondria were measured by using an inhibitor (atractyloside) stop method. Kinetic data of carrier-specific bound analogues were evaluated from Dixon plots and indicate that these analogues are competitive inhibitors for mitochondrial [14C]ADP uptake. Km and Vmax values for carrier-mediated uptake of nucleotide analogues were calculated from Lineweaver-Burk plots. By means of the analogues, a systematic mapping of the essential chemical and steric interactions between the transporter protein and the heterocycle of its substrate in the course of the binding as well as transfer step was achieved. Prerequisites for carrier-specific binding (recognition) are (A) an anti- or syn-positioned beta-glycosyl-linked heterocycle, (B) a nitrogen ring atom in position 7 for syn-structured analogues, and (C) an electron-rich region at the N(1) position, i.e., a permanent dipole moment oriented toward N(1) for anti-structured analogues. Additional requirements for subsequent transport catalysis are (A) a non-fixed anti-positioned base moiety with a beta-glycosyl torsion angle of about -20 degrees, (B) a C(6)-positioned amino group, and (C) an unsubstituted C(2) atom. The complementary binding site at the carrier protein to the N(1)-C(6)(-NH2) amidine region is proposed to be represented by two juxtaposed and invariant bonding points, i.e., an asparagine or glutamine residue.

Adenosine Diphosphate↗
PubMed1978

beta-Nicotinamide-alpha-adenine dinucleotide. Synthesis and properties of a coenzyme analog.

It was not possible to synthesize beta-nicotinamide-alpha-adenine dinucleotide (NalphaAD) via the NAD pyrophosphorylase-catalyzed reaction of beta-nicotinamide mononucleotide and alpha-adenosine-5'-O-triphosphate. It was therefore prepared by reacting a mixture of beta-nicotinamide mononucleotide and alpha-adenosine-5'-O-monophosphate in aqueous pyridine, using N,N'-dicyclohexylcarbodiimide as condensing agent. NalphaAD can replace NAD in the lactate-dehydrogenase-catalyzed oxidation of lactate to pyruvate with LDH 4M from hog muscle as well as LDH 4H from pig heart. Km values are found to be about one order of magnitude higher for NalphaAD than for NAD. The turnover number for NalphaAD with LDH 4H is similar to that of NAD (95%), whereas it is reduced to one third (35%) in the reaction with LDH 4M. Experiments findings are discussed on the lines of a NAD-equivalent conformation of NalphaAD in the holoenzyme complex.

Animals↗
PubMed1976

Studies on the adenine nucleotide translocase from rat liver mitochondria. Isolation, partial characterization and immunochemical properties of carboxyatractylate-binding protein.

1. Solubility of mitochondrial membranes in various solvent systems was determined quantitatively. The most effective agent was the anionic detergent, sodium dodecylsulphate, which solubilizes 90% of the protein at the concentration of 0.1% followed by Triton X-100 (70%), sodium deoxycholate (60%), Brij 56 (50%), and guanidine hydrochloride (40%) at a concentration of 2 M. 2. Affinity chromatography of a clear 0.1% sodium dodecylsulphate solution of digitonized mitochondria on Sepharose 4B containing carboxyatractylate always resulted in the separation of two fractions, one of which was not retained by the column and the other which could be obtained after elution with 2% sodium dodecylsulphate. 3. The retained protein showed a high binding specificity for ATP and [3H]atractylate when compared with the unretained fraction. The amount of bound [3H]atractylate or carboxyatractylate-sensitive binding of ATP was 10.5 +/- 4 nmol/mg protein, and 22 +/- 8 nmol/mg protein, respectively. 4. The major component within the retained fraction, comprising 85% of the total weight, was protein, followed by phospholipids (14%) and approximately 1% triglycerides. Sodium dodecylsulphate-polyacrylamide gel electrophoresis revealed a major (95%) and a minor (5%) component with an apparent molecular weight of 26000 +/- 1000 and 8300 +/- 400, respectively. The gels did not stain for carbohydrates. Ultracentrifugal analysis showed a single, symmetrical boundry. 5. Double immunodiffusion analysis gave a single precipitin line with the corresponding antiserum. [14C]ADP exchange of digitonin particles was completely inhibited by an antiserum to the carboxyatractylate binding protein fraction, whereas the adenine nucleotide transport of intact mitochondria remained unaffected. In the presence of specific immunoglobulins state-3 respiration rate of digitonin particles was prolonged and reduced by approximately 25%. State-4 respiration rate was unaffected.

Adenosine Triphosphate↗
PubMed1976

[Chromatographic investigation of the substrate properties of 8-bromo-ATP in the nucleoside diphosphate kinase reaction (author's transl)].

8-bromo-ATP is a substrate of nucleoside diphosphate kinase. Its reactivity is similar to that of ATP, and the reaction appears to be unaffected by the different conformations of these two substrates. The nucleoside diphosphate kinase reaction with 8-bromo-ATP cannot be monitored photometrically (optical test) using an NADH/NAD+-dependent enzyme system, because the product 8-bromo-ADP is not a substrate for pyruvate kinase. Measurements of the nucleoside diphosphate kinase reaction with 8-bromo-ATP therefore, were carried out by chromatographic analysis of the nucleoside-triphosphate/diphosphate ratio.

Adenosine Triphosphate↗
PubMed1975

Properties of the ribose-ring-opened adenine nucleotide, 2,2'(1-(9-adenyl)-1'-(tri-,diphosphoryl-oxymethyl))-dihydroxydiethylether in mitochondrial adenine-nucleotide translocation.

(Adenine-14C) or (gamma-32P)-labelled 2,2'[1-(9-adenyl)-1'-(tri-, diphosphoryl-oxymethyl]-dihydroxydiethylether (rroANP) was obtained from ANP by cleavage of the C-2--C-3' bond by sodium periodate oxidation and subsequent borohydride reduction. Binding of rroANP to rat liver mitochondria revealed carrier-linked (atractyloside-sensitive) and unspecific (atractyloside-insensitive) binding but no transfer across the inner mitochondrial membrane. Kinetic data indicate rroANP as a competitive inhibitor for ANP uptake with Ki = 9.3 X 10(-5) M. Experimental rroANP confirmed that an intact adenine base and three anio.nic charges of the phosphate chain are essential for the recognition between ANP-carrier and nucleotide but insufficient for the induction of a transmembrane ANP exchange. In addition mobilisation of the carrier-nucleotide complex requires an intact ribofuranoside ring system.

Adenine Nucleotides↗