Biomedical subjects

M Götte

Publications and source records attributed to M Götte.

24 records · Page 2Linked to original sources

PubMed1996

Probing the higher order structure of RNA with peroxonitrous acid.

Potassium peroxonitrite (ONOOK) and [Fe(EDTA)]2- were used to analyze the influence of chemically entirely different hydroxyl radical sources on tRNA cleavage profiles. [Fe(EDTA)]2- gives rise to hydroxyl radicals via a Fenton-like reaction during the oxidation of chelated Fe2+, while ONOOK generates hydroxyl radicals via its conjugate acid (ONOOH) when adding a stable alkaline solution of ONOOK in samples buffered at neutral pH. [Fe(EDTA)]2- is known to induce oxidative strand scission at sugar moieties thought to be solvent accessible, while those residues located in the 'inside' of structured RNAs are protected. Although ONOOH is neutral and significantly smaller than the metal complex, both reagents generate the same protection pattern on tRNAs, suggesting that access of the commonly formed hydroxyl radical, rather than access of its source, is the determining factor when probing the higher order structure of RNA. Strong difference in reactivity is only seen at the modified 2-thiouridine S34 of tRNA(Lys3) which shows hyperreactivity towards ONOOK treatment. This particular reaction may require interaction between the peroxonitrite anion and the thiocarbonyl group of the base, since hyperreactivity is not observed when probing the dethiolated tRNA(Lys3).

Animals↗

PubMed1995

HIV-1 reverse transcriptase-associated RNase H cleaves RNA/RNA in arrested complexes: implications for the mechanism by which RNase H discriminates between RNA/RNA and RNA/DNA.

Reverse transcription of human immunodeficiency virus type 1 (HIV-1) is primed by tRNA(Lys3), which forms an 18 base pair RNA homoduplex with its 3' terminus and the primer binding site (PBS) of the viral genome. Using an in vitro system mimicking initiation of minus strand DNA synthesis, we analyzed the mechanism by which HIV-1 reverse transcriptase (RT)-associated ribonuclease H (RNase H) distinguishes between RNA/DNA and RNA/RNA (dsRNA). tRNA(Lys3) was hybridized to a PBS-containing RNA template and extended by addition of deoxynucleoside triphosphates (dNTPs). In the presence of all four dNTPs, initial cleavage of the RNA template occurred immediately downstream of the tRNA-DNA junction, reflecting RNase H specificity for RNA in a RNA/DNA hybrid. However, in the absence of DNA synthesis, or limiting this by chain termination, the PBS was cleaved at a constant distance of 18 nucleotides upstream of the nascent primer 3' terminus. The position of cleavage remained in register with the position of DNA synthesis arrest, indicating that hydrolysis of homoduplex RNA is spatialy co-ordinated with DNA synthesis. Kinetic studies comparing cleavage rates of an analogous DNA primer/PBS heteroduplex and the tRNA(Lys3)/PBS homoduplex showed that while the former is cleaved as rapidly as RT polymerizes, the latter proceeds 30-fold slower. Although the RNase H domain hydrolyzes dsRNA when RT is artificially arrested, specificity for RNA/DNA hybrids is maintained when DNA is actively synthesized, since residency of the RNase H domain at a single base position is not long enough to allow significant cleavage on dsRNA.

Base Sequence↗

PubMed1995

Endocytosis of decorin by bovine aortic endothelial cells. off.

The small dermatan sulfate proteoglycan decorin is efficiently internalized by a variety of cells of mesenchymal origin. Decorin-binding receptor proteins of 51 and 26 kDa are involved in this process. Uptake is modulated by highly sulfated heparan sulfate which interacts with the receptor proteins, too. Compared with cultured skin fibroblasts, bovine aortic endothelial cells have a lower capacity for decorin endocytosis whereas their apparent concentration of receptor proteins is even higher. The low internalization rate is attributed to the greater occupancy of receptor proteins by heparan sulfate of the plasma membrane and/or the extracellular matrix. Growth of endothelial cells on Falcon 3090 tissue culture inserts made possible to study decorin uptake from the apical and basolateral membrane, respectively. Decorin uptake was at the limit of detection when the proteoglycan was added to the basolateral compartment. Uptake via the apical membrane was at least as efficient as in monolayer cultures on plastic. The basolateral membrane, however, was enriched in receptor proteins, but also in heparan sulfate proteoglycans. It is, therefore, suggested that endothelial cells are especially involved in the clearance of decorin which is present in blood plasma.

Animals↗

PubMed1994

The 'helix clamp' in HIV-1 reverse transcriptase: a new nucleic acid binding motif common in nucleic acid polymerases.

Amino acid sequences homologous to 259KLVGKL (X)16KLLR284 of human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) are conserved in several nucleotide polymerizing enzymes. This amino acid motif has been identified in the crystal structure model as an element of the enzyme's nucleic acid binding apparatus. It is part of the helix-turn-helix structure, alpha H-turn-alpha I, within the 'thumb' region of HIV-1 RT. The motif grasps the complexed nucleic acid at one side. Molecular modeling studies on HIV-1 RT in complex with a nucleic acid fragment suggest that the motif has binding function in the p66 subunit as well as in the p51 subunit, acting as a kind of 'helix clamp'. Given its wide distribution within the nucleic acid polymerases, the helix clamp motif is assumed to be a structure of general significance for nucleic acid binding.

Algorithms↗

PubMed1991

A novel large dermatan sulfate proteoglycan from human fibroblasts.

Human skin fibroblasts express, in addition to versican, a second large chondroitin sulfate/dermatan sulfate proteoglycan, which has been investigated with the aid of a specific antiserum in cultures of fetal fibroblasts. Its core protein, obtained after chondroitin ABC lyase treatment, exhibits an apparent molecular mass of about 740 kDa in the absence of a reducing agent whereas reduction produces two core proteins of 460 and 300 kDa, respectively. Both subunits carry one or very few dermatan sulfate chains of about 20 kDa which are of similar chemical composition irrespective of the type of subunits to which they are attached. Tryptic peptide maps of [35S]methionine-labeled core proteins indicated that both subunits are related neither to each other nor to versican, suggesting that the proteoglycan exists predominantly as a heterodimeric molecule. It is insensitive to collagenase and does not interact with hyaluronan. Pulse-chase experiments suggested that the core proteins are different gene products. Dimerization begins soon after core protein synthesis but requires more than 2 h for completion. Glycosaminoglycan synthesis occurs immediately prior to secretion. A small proportion of both subunits may be secreted in form of a monomeric proteoglycan. The heterodimeric proteoglycan is a major proteoglycan species of fetal fibroblasts. The secreted product represents 10-20% of [35S]methionine and about 5-10% of [35S]sulfate incorporated into secreted proteoglycans.

Cells, Cultured↗

PubMed1985

[The increase in the liberation rate of propyphenazone from preoral solid dosage forms].

The authors report on the influence of the granulation and tabletting applying macromolecular agents (polyvinylpolypyrrolidone Heweten 40, polyethylenglycol 6000, polyvinylpyrrolidone K25, polyvinylpyrrolidone K90 and potato starch) on the liberation rate of propyphenazone which exhibits by its hydrophobicity unfavourable dissolution parameters. A significant increase of the liberation rate of propyphenazone (re-crystallized from various media) has been realized by direct pressing of polyvinylpolypyrrolidone and Heweten 40 as well as by gelatine solution granulation, with which the last method exhibits the advantage of an increased accuracy of dosage and a decreased friability. The gelatine granulation of the drug itself as well as the starch granulate addition resulted in an increased dissolution rate. The addition of other agents (polyvinylpyrrolidone K25, K90 and polyethylenglycol 6000) as well as the pressing of a propyphenazone produced by re-crystallization of an aqueous tenside solution, which distinguishes by an extreme dissolution rate, were not at all suitable for the tabletting. The tablet disintegration caused by hydrophile disintegration media, is the condition granting the liberation of the hydrophobic drug. The wettability and thus the dissolution rate of the drug are increased by the liberation.

Antipyrine↗