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Biomedical subjects

T Schindler

Publications and source records attributed to T Schindler.

At least 19 recordsLinked to original sources
PubMed2006

[Palliative care in Germany].

The delivery of palliative care in Germany is still characterized by a wide-spread undersupply both for inpatients and outpatients. Nevertheless over the last 15 years progress has also been made in Germany, which is pleasing and which has directed professional and public attention increasingly to the situation of the incurably ill and dying. In the course of this development the first structures for specialized palliative care have been established. In particular the situation for inpatients in hospitals and hospices is pleasing, even if a satisfaction for total coverage is not yet reached. More seriously however is the demand for the outpatients. Financial means, which are steered so far only rudimentarily for palliative care, must be made sufficiently available, in order to be able to meet the demand with the help of appropriate structural offers. It is important to realize the needs of the incurably ill and dying and of their relatives and friends. Our society has to call attention to this subject and it should be placed on the healthpolitical agenda as a topic of the highest priority level. The current developments in the federation and in the federal states are encouraging and allowing hope that the delivery of palliative care for all who need it (and not only the ill) will improve further in the coming years.

Germany↗
PubMed2003

[Appropriate uptake period for myocardial PET imaging with 18F-FDG after oral glucose loading].

AIM: Identification of a rationale for the appropriate uptake period for myocardial (18)F-FDG-PET imaging of patients with and without diabetes mellitus. METHODS: In a subset of 27 patients, static 2D-PET examination was performed of patients with chronic coronary artery disease and known myocardial infarction. The patients fasted (at least 4 h) before examination. (18)F-FDG (330 +/- 20 MBq) was injected intravenously. The image quality was semiquantitativly determined by ROI-analysis and the myocardium-to-blood pool activity ratio (M/B) was calculated. I.) Scans 30, 60, and 90 min p. i. of 10 non-diabetic patients (60 g oral glucose loading one hour before FDG-injection, low-dose intravenous insulin bolus if necessary). II.) Scans 30, 60, and 90 min p. i. of 10 patients with known non-insulin dependent diabetes (20 g glucose, insulin bolus). III.) Scans 90 min p. i. of 7 patients with known non-insulin dependent diabetes and elevated fasting serum glucose level (140-200 mg/dl; insulin bolus, no glucose). RESULTS: I.) The M/B ratio significantly increases in nondiabetic patients with the uptake time (30 min 1.95 +/- 0.20; 60 min 2.96 +/- 0.36; 90 min 3.78 +/- 0.43). II.) In patients with non-insulin dependent diabetes the M/B ratio also significantly increases with uptake time. Compared to non-diabetic patients group II reached smaller M/B values (30 min 1.56 +/- 0.10; 60 min 2.15 +/- 0.14; 90 min 2.71 +/- 0.19). III.) In the group of patients with elevated fasting serum glucose level (who only got insulin but no glucose loading) the M/B activity ratio 90 min p. i. was clearly inferior compared with diabetic patients after oral glucose loading and insulin administration (M/B 2.71 +/- 0.19 versus 2.16 +/- 0.07). CONCLUSIONS: In static myocardial viability PET studies with (18)F-FDG an uptake time of 90 min yields image quality superior to that obtained after shorter uptake time.

Aged↗
PubMed2001

Phosphorylation and structure-based functional studies reveal a positive and a negative role for the activation loop of the c-Abl tyrosine kinase.

c-Abl is a nuclear and cytoplasmic tyrosine kinase involved in a variety of cellular growth and differentiation processes. In contrast to its oncogenic counterparts, like BCR-Abl, c-Abl is not constitutively tyrosine phosphorylated and its catalytic activity is very low. Here we report tyrosine phosphorylation of endogenous c-Abl and a concomitant increase in catalytic activity. Using Abl -/- cells reconstituted with mutated c-Abl forms, we show that phosphorylation and activity depend on Tyr412 in the activation loop. Tyr412 is also required for stimulation by PDGF or by cotransfection of active Src. Phosphorylation of Tyr412 can occur autocatalytically by a trans-mechanism and cause activation of otherwise inactive c-Abl, suggesting a positive feedback loop on c-Abl activity. In the recent structure of the Abl catalytic domain bound to the STI-571 inhibitor, unphosphorylated Tyr412 in the activation loop points inward and appears to interfere with catalysis. We mutated residues involved in stabilizing this inhibited form of the activation loop and in positioning Tyr412. These mutations resulted in tyrosine phosphorylation and activation of c-Abl, as if relieving c-Abl from inhibition. Tyr412 is therefore necessary both for activity and for regulation of c-Abl, by stabilizing the inactive or the active conformation of the enzyme in a phosphorylation-dependent manner.

Blotting, Western↗
PubMed2001

DcAGP1, a secreted arabinogalactan protein, is related to a family of basic proline-rich proteins.

A cDNA corresponding to the core protein of an immunoaffinity-purified arabinogalactan protein (AGP) secreted aucus carota (carrot) cells in liquid culture was isolated. This cDNA, DcAGP1, encodes a new class of non-classical' AGP with strong similarity to a family of basic proline-rich proteins. The protein is rich in proline (17%), alanine (10%) and lysine (11%) and contains four distinct domains: a signal peptide, a proline-rich domain, a histidine-rich basic domain and a cysteine-containing 'PAC' domain that is found in a range of other cell wall proteins. The protein contains several sequence motifs found in otherwise unrelated cell wall proteins, but also displays some unique features. Northern blot analyses show that while the DcAGP1 transcript is abundant in the suspension-culture cells from which the AGP was obtained; in carrot seedlings the gene is only expressed at low levels in the roots and it is neither wound- nor stress-inducible. Furthermore, northern and western blot analyses demonstrate that the core polypeptide of DcAGP1 is differentially glycosylated in two different carrot suspension cultures. The unusual features of the protein sequence suggest that the DcAGP1 protein is a member of a family of basic proline-rich proteins defined by the C-terminal PAC domain, and the possible function(s) of the DcAGP1 protein is considered in the light of current views on AGP structure and function.

Amino Acid Sequence↗
PubMed2000

Structural mechanism for STI-571 inhibition of abelson tyrosine kinase.

The inadvertent activation of the Abelson tyrosine kinase (Abl) causes chronic myelogenous leukemia (CML). A small-molecule inhibitor of Abl (STI-571) is effective in the treatment of CML. We report the crystal structure of the catalytic domain of Abl, complexed to a variant of STI-571. Critical to the binding of STI-571 is the adoption by the kinase of an inactive conformation, in which a centrally located "activation loop" is not phosphorylated. The conformation of this loop is distinct from that in active protein kinases, as well as in the inactive form of the closely related Src kinases. These results suggest that compounds that exploit the distinctive inactivation mechanisms of individual protein kinases can achieve both high affinity and high specificity.

Animals↗
PubMed2000

Reciprocal regulation of Hck activity by phosphorylation of Tyr(527) and Tyr(416). Effect of introducing a high affinity intramolecular SH2 ligand.

The Src family tyrosine kinase Hck possesses two phosphorylation sites, Tyr(527) and Tyr(416), that affect the catalytic activity in opposite ways. When phosphorylated, Tyr(527) and residues C-terminal to it are involved in an inhibitory intramolecular interaction with the SH2 domain. However, this sequence does not conform to the sequence of the high affinity SH2 ligand, pYEEI. We mutated this sequence to YEEI and show that this mutant form of Hck cannot be activated by exogenous SH2 ligands. The SH3 domain of Hck is also involved in an inhibitory interaction with the catalytic domain. The SH3 ligand Nef binds to and activates YEEI-Hck mutant in a similar manner to wild-type Hck, indicating that disrupting the SH3 interaction overrides the strengthened SH2 interaction. The other phosphorylation site, Tyr(416), is the autophosphorylation site in the activation loop. Phosphorylation of Tyr(416) is required for Hck activation. We mutated this residue to alanine and characterized its catalytic activity. The Y416A mutant shows a higher K(m) value for peptide and a lower V(max) than autophosphorylated wild-type Hck. We also present evidence for cross-talk between the activation loop and the intramolecular binding of the SH2 and SH3 domains.

Amino Acid Sequence↗
PubMed1999

Microsecond folding of the cold shock protein measured by a pressure-jump technique.

A pressure-jump apparatus was employed in investigating the kinetics of protein unfolding and refolding. In the reaction cell, the pressure can be increased or decreased by 100-160 bar within 50-100 microseconds and then held constant. Thus, unfolding and refolding reactions in the time range from 70 microseconds to 70 s can be followed with this technique. Measurements are possible in the transition regions of thermally or denaturant-induced folding in a wide range of temperatures and solvent conditions. We used this pressure-jump method to determine the temperature dependence of the rate constants of unfolding and refolding of the cold shock protein of Bacillus subtilis and of three variants thereof with Phe --> Ala substitutions in the central beta-sheet region. For all variants, the change in heat capacity occurred in refolding between the unfolded and activated states, suggesting that the overall native-like character of the activated state of folding was not changed by the deletion of individual Phe side chains. The Phe27Ala mutation affected the rate of unfolding only; the Phe15Ala and Phe17Ala mutations changed the kinetics of both unfolding and refolding. Although the activated state of folding of the cold shock protein is overall native-like, individual side chains are still in a non-native environment.

Alanine↗
PubMed1999

The family of cold shock proteins of Bacillus subtilis. Stability and dynamics in vitro and in vivo.

Bacillus subtilis possesses three homologous small cold shock proteins (CSPs; CspB, CspC, CspD, sequence identity >72%). They share a similar beta-sheet structure, as shown by circular dichroism, and have a very low conformational stability, with CspC being the least stable. Similar to CspB, CspC and CspD unfold and refold extremely fast in a N <==> U two-state reaction with average lifetimes of only 100-150 ms for the native state and 1-6 ms for the unfolded states at 25 degreesC. As a consequence of their low stability and low kinetic protection against unfolding, all three cold shock proteins are rapidly degraded by proteases in vitro. Analysis of the CSP stabilities in vivo by pulse-chase experiments revealed that CspB and CspD are stable during logarithmic growth at 37 degreesC as well as after cold shock. The cellular half-life of CspC is shortened at 37 degreesC, but under cold shock conditions CspC becomes stable. The proteolytic susceptibility of the CSPs in vitro was strongly reduced in the presence of a nucleic acid ligand, suggesting that the observed stabilization of CSPs in vivo is mediated by binding to their substrate mRNA at 37 degreesC and, in particular, under cold shock conditions.

Amino Acid Sequence↗
PubMed1999

A protein folding intermediate of ribonuclease T1 characterized at high resolution by 1D and 2D real-time NMR spectroscopy.

The rate-limiting step during the refolding of S54G/P55N ribonuclease T1 is determined by the slow trans-->cis prolyl isomerisation of Pro39. We investigated the refolding of this variant by one-dimensional (1D) and two-dimensional (2D) real-time NMR spectroscopy, initiated by a tenfold dilution from 6 M guanidine hydrochloride at 10 degreesC. Two intermediates could be resolved with the 1D approach. The minor intermediate, which is only present early during refolding, is largely unfolded. The major intermediate, with an incorrect trans Pro39 peptide bond, is highly structured with 33 amide protons showing native chemical shifts and native NOE patterns. They could be assigned in a real-time 2D-NOESY (nuclear Overhauser enhancement spectroscopy) by using a new assignment strategy to generate positive and negative signal intensities for native and non-native NOE cross-peaks, respectively. Surprisingly, amide protons with non-native environments are located not only close to Tyr38-Pro39, but are spread throughout the entire protein, including the C-terminal part of the alpha-helix, beta-strands 3 and 4 and several loop regions. Native secondary and tertiary structure was found for the major intermediate in the N-terminal beta-strands 1 and 2 and the C terminus (connected by the disulfide bonds), the N-terminal part of the alpha-helix, and the loops between beta-strands 4/5 and 5/6. Implications of these native and non-native structure elements of the intermediate for the refolding of S54G/P55N ribonuclease T1 and for cis/trans isomerizations are discussed.

Magnetic Resonance Spectroscopy↗
PubMed1999

Crystal structure of Hck in complex with a Src family-selective tyrosine kinase inhibitor.

The crystal structure of the autoinhibited form of Hck has been determined at 2.0 A resolution, in complex with a specific pyrazolo pyrimidine-type inhibitor, PP1. The activation segment, a key regulatory component of the catalytic domain, is unphosphorylated and is visualized in its entirety. Tyr-416, the site of activating autophosphorylation in the Src family kinases, is positioned such that access to the catalytic machinery is blocked. PP1 is bound at the ATP-binding site of the kinase, and a methylphenyl group on PP1 is inserted into an adjacent hydrophobic pocket. The enlargement of this pocket in autoinhibited Src kinases suggests a route toward the development of inhibitors that are specific for the inactive forms of these proteins.

Animals↗
PubMed1999

Vitamin C improves endothelial function of epicardial coronary arteries in patients with hypercholesterolaemia or essential hypertension--assessed by cold pressor testing.

AIMS: There is evidence that formation of free radicals increases in patients with hypertension or hypercholesterolaemia, which may contribute to endothelial dysfunction of epicardial coronary arteries due to inactivation of the vasodilator NO. The present study was designed to test whether the abnormal constriction of epicardial coronary arteries due to sympathetic stimulation by the cold pressor test in patients with essential hypertension or hypercholesterolaemia could be reversed by administration of the antioxidant vitamin C. METHODS AND RESULTS: In 28 patients without relevant coronary artery stenosis the cold pressor test was performed before and after a 3 g infusion of vitamin C. In five normal controls the cold pressor test led to a similar increase in luminal area before and after vitamin C (3.7+/-1.3% and 1.9+/-0.8%, ns vs before vitamin C). In nine hypercholesterolaemic patients the cold pressor test led to a -14.1+/-2.8% reduction in cross-sectional area before vitamin C. This constriction was significantly improved after vitamin C to -7.6%+/-2.0, P=0.027 vs before vitamin C. In nine hypertensive patients, the cold pressor test led to a -17.1+/-3.2% decrease in cross-sectional area before vitamin C, which was improved to -7.1+/-3.1 after vitamin C, P=0.004 vs before vitamin C. This increase in luminal area was significant in each group in comparison with normal controls (each P<0.05). Administration of saline (placebo group, five patients) had no significant effect on cold pressor test-induced constriction (-6.9+/-3.9% before and -6. 8+/-3.7% after saline). CONCLUSION: The antioxidant vitamin C reverses cold pressor test-induced vasoconstriction of epicardial coronary arteries in patients with hypertension or hypercholesterolaemia. Our data suggest that enhanced oxidative stress contributes to impaired endothelial function in this patient population.

Aged↗
PubMed1998

Surface-exposed phenylalanines in the RNP1/RNP2 motif stabilize the cold-shock protein CspB from Bacillus subtilis.

In the cold-shock protein CspB from Bacillus subtilis three exposed Phe residues (F15, F17, and F27) are essential for its function in binding to single-stranded nucleic acids. Usually, the hydrophobic Phe side chains are buried in folded proteins. We asked here whether the exposition of the essential Phe residues could be a cause for the very low conformational stability of CspB. Urea-induced and heat-induced equilibrium unfolding transitions were measured for three mutants of CspB, where Phe 15, Phe 17, and Phe 27 were individually replaced by alanine. Unexpectedly, all three mutations strongly destabilized CspB. The aromatic side chains of Phe 15, Phe 17, and Phe 27 in the active site are thus important for both binding to nucleic acids and conformational stability. There is no compromise between function and stability in the active site. Model calculations indicate that, although they are partially exposed to solvent, all three Phe residues nevertheless lose accessible surface upon folding, and this should favor the native state. A different result is obtained with the F38A variant. Phe 38 is hyperexposed in native CspB, and its substitution by Ala is in fact stabilizing.

Bacillus subtilis↗
PubMed1998

Intracortical functional heterogeneity in area striate during penicillin-induced spikes in rabbits.

The generation and spread of epileptiform activity within the cortex depend on the functional and anatomical relationships between the focus and its surrounding area. These processes are not completely understood. Thus intracortical current-source-density analysis (CSD) was performed in six rabbits in order to investigate this functional relationship. Electric potential was measured perpendicular to the cortical surface by means of two 16-channel probes, and CSD was calculated within the focus and at various distances of up to 5 mm. The cortical areas surrounding the focus could be subdivided into three regions. The region up to 3 mm from the focus showed similar activity but beyond 4.5 mm no characteristic functional relationship was found with regard to the epileptiform events within the focus. Within the region 3.5-4.5 mm, however, mainly supragranular cells seem to contribute to the electric potential measured at the cortical surface and within the extracellular space. They were activated simultaneously with the initiation of focal spike generation. Taking into account the distribution of the electric potential and the results of CSD analysis, these cells seem mainly involved in the inhibition of the horizontal spread of spike activity.

Action Potentials↗
PubMed1998

Conservation of rapid two-state folding in mesophilic, thermophilic and hyperthermophilic cold shock proteins.

The cold shock protein CspB from Bacillus subtilis is only marginally stable, but it folds extremely fast in a simple N reversible U two-state reaction. The corresponding cold shock proteins from the thermophile Bacillus caldolyticus and the hyperthermophile Thermotoga maritima show strongly increased conformational stabilities, but unchanged very fast two-state refolding kinetics. The absence of intermediates in the folding of B. subtilis CspB is thus not a corollary of its low stability. Rather, two-state folding and an unusually native-like activated state of folding seem to be inherent properties of these small all-beta proteins. There is no link between stability and folding rate, and numerous sequence positions exist which can be varied to modulate the stability without affecting the rate and mechanism of folding.

Amino Acid Sequence↗
PubMed1998

[Value of color-coded duplex ultrasound in evaluating arm blood vessels--arteries and hemodialysis shunts].

AIM: Comparison of colour coded duplex sonography (CCDS) to angiography in visualizing arteries and haemodialysis fistulas of the upper extremity. METHOD: 1145 arterial and vein segments were visualised using CCDS and angiography in 219 patients by different examiners with different degrees of experience. Sensitivity, specificity and accuracy were calculated. RESULTS: For the arterial segments--subclavian artery up to the distal lower arm arteries--the sensitivity was 90% and the specificity 99%. The sensitivity for the detection of vascular alterations in haemodialysis fistulas was 89% and the specificity 95%. CONCLUSION: CCDS may be able to replace diagnostic angiography of the arm except when evaluating the hand arteries and the central veins.

Adult↗