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G Müller

Publications and source records attributed to G Müller.

At least 289 records · Page 16Linked to original sources
PubMed1994

Lipolytic membrane release of two phosphatidylinositol-anchored cAMP receptor proteins in yeast alters their ligand-binding parameters.

Two new cAMP-binding proteins have been discovered recently in Saccharomyces cerevisiae. They are genetically distinct from the regulatory subunit of cytoplasmic cAMP-dependent protein kinase A and are distinguished from the latter, in addition, by their anchorage through phosphatidylinositol-containing lipid and glycolipid structures to mitochondrial and plasma membranes, respectively (Müller and Bandlow, 1989 Biochemistry 28, 9957-9967, 1991, Biochemistry 30, 10181-10190). A nutritional upshift induces the cleavage of the anchor by a phospholipase C (Müller and Bandlow, 1993, J. Cell Biol. 122, 225-236). To test the idea that anchorage by (glycosyl)phosphatidyl-inositol influences cAMP-binding and has a regulatory function, we analyzed ligand binding to the two purified cAMP receptors (46,000 and 54,000 Da) in comparison to the regulatory subunit of the cytoplasmic protein kinase A (52,000 Da). We find that lipolytic cleavage of the two membrane anchors by phosphatidylinositol-specific phospholipases C and D results in significantly higher association and lower dissociation rates of cAMP, thus leading to a dramatic increase in ligand affinity of the two cAMP receptors. Use of cAMP analogues identifies two different cAMP-binding centers in each membrane-embedded protein, one of which is noticeably affected by the cleavage of the anchor. In both phosphatidylinositol-anchored cAMP receptor proteins a single Trp residue in one of the binding centers is photoaffinity-labeled by 8-N3-cAMP, whereas two amino acids, Trp and Tyr, are modified after lipolytic removal of the anchor. The differences in the labeling patterns are interpreted as to result from a conformational rearrangement induced by the cleavage of the anchor. Together with the increased affinity to the ligand these changes document alterations of the properties and folding structure of lipid-anchored proteins following cleavage of the PI-containing anchor by specific phospholipases and provide the first molecular evidence for a regulatory role of the anchorage by a lipid structure. The cytoplasmic regulatory subunit of yeast protein kinase A is not photolabeled to a significant extent under any condition.

Affinity Labels↗
PubMed1994

Cloning and expression of three isoforms of the human EP3 prostanoid receptor.

Functional cDNA clones coding for three isoforms of the human prostaglandin E receptor EP3 subtype have been isolated from kidney and uterus cDNA libraries. The three isoforms, designated hEP3-I, hEP3-II and hEP3-III, have open reading frames corresponding to 390, 388 and 365 amino acids, respectively. They differ only in the length and amino acid composition of their carboxy-terminal regions, beginning at position 360. The human EP3 receptor has seven predicted transmembrane spanning domains and therefore belongs to the G-protein-coupled receptor family. The rank order of potency for prostaglandins and related analogs in competition for [3H]PGE2 specific binding to membranes prepared from transfected COS cells was comparable for all three isoforms, and as predicted for the EP3 receptor, with PGE2 = PGE1 >> PGF2 alpha = iloprost > PGD2 >> U46619. In addition, the EP3-selective agonist MB28767 was a potent competing ligand with an IC50 value of 0.3 nM, whereas the EP1-selective antagonist AH6909 gave IC50 values of 2-7 microM and the EP2-selective agonist butaprost was inactive. In summary, we have cloned three isoforms of the human EP3 receptor having comparable ligand binding properties.

Amino Acid Sequence↗
PubMed1994

Porin proteins in mitochondria from rat pancreatic islet cells and white adipocytes: identification and regulation of hexokinase binding by the sulfonylurea glimepiride.

The binding of hexo-/glucokinase and glycerol kinase to mitochondria via the channel forming protein, porin, in pancreatic islet beta-cells and adipocytes, was recently proposed to participate in nutritional signaling, glucose sensing, and the control of high-energy phosphate distribution and oxidative phosphorylation. In this study we demonstrate that polyclonal antisera against purified rat liver porin recognize unique proteins in rat pancreatic islets, adipocytes, and RINm5F cells, each with an apparent M(r) about 2000 smaller than that of liver porin. Immunoblotting of subcellular fractions, the purity of which has been controlled by the distribution of marker proteins, revealed the mitochondrial localization of the cross-reacting proteins. Their enrichment with a method used for the purification of porin proteins, the characteristic behavior during isoelectric focusing, and the specific binding of rat liver hexokinase and glycerol kinase to phospholipid vesicles containing the purified cross-reacting beta-cell or adipocyte proteins strongly suggest their identity with mitochondrial porin. The subtle differences in the apparent M(r) and charge heterogeneity raise the possibility of the existence of porin isoforms expressed in a tissue-specific manner. Anti-porin antisera coimmunoprecipitated hexo-/glucokinase from rat insulinoma cell (RINm5F) and adipocyte mitochondria as determined by subsequent immunoblotting of the immunoprecipitates with polyclonal antisera against yeast hexokinase and rat liver glucokinase, respectively. This indicates that some rat pancreatic glucokinase (54 kDa) and liver hexokinase (102 kDa), respectively, is bound to mitochondrial porin. The major portion of the bound fraction is released from mitochondria after treatment with glucose 6-phosphate. Incubation of RINm5F and fat cells with the insulin releasing sulfonylurea drug, glimepiride (20 nM and 5 microM, respectively) for 30 min reduces the amount of hexo-/glucokinase associated with mitochondria and porin to about 50-30%. The reduced kinase binding activity of porin is preserved after isolation of porin from glimepiride-treated cells, reconstitution into phospholipid vesicles and assaying for glucose 6-phosphate inhibitable binding of rat liver hexokinase. The sulfonylurea tolbutamide (20 microM and 5 mM) is significantly less effective. The sulfonylurea-induced inhibition of hexo-/glucokinase binding to mitochondrial porin does not require glucose metabolism or Ca2+ influx into the cells. These data suggest that the sulfonylurea glimepiride, which is thought to inhibit the ATP-regulated K(+)-channel in beta-cells, may have, in addition, an intracellular site of action in pancreatic islet and adipocyte cells at the level of regulation of gluco-/hexokinase binding to mitochondrial porin.

Adipocytes↗
PubMed1994

4'-Amino-benzamido-taurocholic acid selectively solubilizes glycosyl-phosphatidylinositol-anchored membrane proteins and improves lipolytic cleavage of their membrane anchors by specific phospholipases.

Glycosyl-phosphatidylinositol-anchored membrane proteins (GPI-proteins) are normally identified either by cleavage of the lipid anchor using (glycosyl)phosphatidylinositol-specific phospholipases C or D (GPI-PLs) or by metabolic labeling of the lipid moiety with specific building blocks. Therefore, methods for discrimination between transmembrane proteins and GPI-proteins on the basis of their physicochemical properties are desirable. Here we are presenting a selective extraction method for typical well-characterized mammalian GPI-proteins, e.g., acetylcholine esterase, alkaline phosphatase, 5'-nucleotidase, and lipoprotein lipase, using a derivative of taurocholate. The results were compared to those obtained with well-characterized transmembrane proteins, e.g., insulin receptor and hydroxymethyl glutaryl coenzyme A-reductase, glucose transporters, or aminopeptidase M and several commercially available detergents. With regard to total membrane proteins, it was possible to selectively enrich GPI-proteins up to 8- to 14-fold by using concentrations between 0.1 and 0.3% of 4'-NH2-amino-7 beta-benzamido-taurocholic acid (BATC). In addition, the cleavage specificity and efficiency of (G)PI-PLs were increased in the presence of identical concentrations of BATC compared to commonly used detergents, e.g., Nonidet P-40. Therefore, the present study shows that the use of BATC facilitates the identification of glycosyl-phosphatidylinositol-anchored membrane proteins.

5'-Nucleotidase↗
PubMed1994

Pharmacophore refinement of gpIIb/IIIa antagonists based on comparative studies of antiadhesive cyclic and acyclic RGD peptides.

Structurally guided design approaches to low-molecular-weight platelet aggregation antagonists addressing the platelet-associated heterodimeric cell surface receptor gpIIb/IIIa rely on comparative studies of an ensemble of conformationally and biologically characterized compounds, since no high-resolution structure of the receptor system is available. We report a classical indirect and comparative pharmacophore refinement approach based on a series of small cyclic Arg-Gly-Asp (RGD) peptides as gpIIb/IIIa-fibrinogen interaction antagonists. These peptides have previously been investigated as potent and selective tumor cell adhesion inhibitors. The definition of geometrical descriptors classifying the RGD peptide conformations and their subsequent analysis over selected RGD- and RXD-containing protein structures allows for a correlation of distinct structural features for platelet aggregation inhibition. An almost parallel alignment of the Arg and Asp side chains was identified by a vector analysis as being present in all active cyclic hexa- and pentapeptides. This orientation is induced mainly by the constraint of backbone cyclization and is not of any covalent tripeptide-inherent origin, which was rationalized by a 500 ps high-energy MD simulation of a sequentially related linear model peptide. The incorporation of the recognition tripeptide Arg-Gly-Asp into the cyclic peptide templates acted as a filter mechanism, restricting the otherwise free torsional relation of both side chains to a parallel orientation. Based on the derived results, several detailed features of the receptor binding site could be deduced in terms of receptor complementarity. These findings should govern the design of next-generation compounds with enhanced activities. Furthermore, the complementary stereochemical characteristics of the substrate can be used as boundary conditions for pseudoreceptor modelling studies that are capable of reconstructing a hypothetical binding pocket, qualitatively resembling the steric and electronic demands of gpIIb/IIIa. It is interesting to note that these features provide clear differentiation to requirements for inhibition of alpha v beta 3 substrate binding. This can account for the extremely high selectivity and activity of some of our constrained peptides for either the alpha IIb beta 3 or the alpha v beta 3 receptor.

Amino Acid Sequence↗
PubMed1994

[PTFE prostheses in infrarenal aneurysms of the abdominal aorta].

Postoperative dilatation of Dacron vascular grafts can lead to aneurysms and rupture of the prosthesis making it necessary to change the graft by operation. Therefore, 15 patients at Lübeck University Hospital underwent replacement of the aorta with PTFE prostheses due to infrarenal aortic aneurysms. Ultrasound measurements 2-12 months postoperatively revealed a dilatation of 11.6% concerning tubular grafts, 11.5% for the shafts and 14.6% for the limbs of bifurcation grafts compared to the higher dilatation rate of Dacron prostheses described in the literature. There was no evidence of anastomotic false aneurysms, thrombosis of the limbs of bifurcation grafts, peripheral embolization or perigraft reaction. In conclusion, PTFE prostheses seem to be a means of preventing patients from having secondary complications as a result of graft dilatation. Based on our early results and the results described in the literature, PTFE prostheses can be regarded as an alternative to dacron prostheses.

Aged↗
PubMed1994

DNA-protein cross-links and sister chromatid exchange frequencies in lymphocytes and hydroxyethyl mercapturic acid in urine of ethylene oxide-exposed hospital workers.

The lymphocytes of 25 hospital workers exposed to ethylene oxide and of a standardized control group were investigated for DNA damage (measured by alkaline filter elution) and sister chromatid exchange (SCE) frequencies. Additionally, the excretion of hydroxyethyl mercapturic acid (HEMA) in the 24-h urine of ten workers and ten control persons was determined. The peak levels of ethylene oxide in air during the first 8 min after opening of the sterilization unit were measured. Peak levels of ethylene oxide in the air of up to 417 ppm after opening of the sterilization unit were detected. In the alkaline filter elution assay we found significantly reduced elution rates in the exposed workers, indicating DNA-protein cross-links. The reduction of the elution rates through HVLP filters correlated significantly with the exposure classification (low, medium, high) (r = -0.45, P < 0.05) and the ethylene oxide peak level after opening of the sterilization unit (r = -0.42, P < 0.05). The SCE frequencies in the standardized control group were significantly elevated. With respect to (n = 78) historic control SCE values of our institute, the SCE values of the disinfectors were not significantly elevated (6.54 vs 6.27). The ethylene oxide-exposed workers did not have a greater percentage of high-frequency SCE cells. The mean HEMA concentration in the urine of the exposed workers was significantly elevated, but there were wide variations in HEMA concentrations and no correlation to ethylene oxide exposure. We conclude that the alkaline filter elution assay may be a sensitive parameter for ethylene oxide-exposed workers.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcysteine↗
PubMed1994

Concentrations of benzene in blood and S-phenylmercapturic and t,t-muconic acid in urine in car mechanics.

Different parameters of biological monitoring were applied to 26 benzene-exposed car mechanics. Twenty car mechanics worked in a work environment with probably high benzene exposures (exposed workers); six car mechanics primarily involved in work organization were classified as non-exposed. The maximum air benzene concentration at the work places of exposed mechanics was 13 mg/m3 (mean 2.6 mg/m3). Elevated benzene exposure was associated with job tasks involving work on fuel injections, petrol tanks, cylinder blocks, gasoline pipes, fuel filters, fuel pumps and valves. The mean blood benzene level in the exposed workers was 3.3 micrograms/l (range 0.7-13.6 micrograms/l). Phenol proved to be an inadequate monitoring parameter within the exposure ranges investigated. The muconic and S-phenylmercapturic acid concentrations in urine showed a marked increase during the work shift. Both also showed significant correlations with benzene concentrations in air or in blood. The best correlations between the benzene air level and the mercapturic and muconic acid concentrations in urine were found at the end of the work shift (phenylmercapturic acid concentration: r = 0.81, P < 0.0001; muconic acid concentration: r = 0.54, P < 0.05). In conclusion, the concentrations of benzene in blood and mercapturic and muconic acid in urine proved to be good parameters for monitoring benzene exposure at the workplace even at benzene air levels below the current exposure limits. Today working as a car mechanic seems to be one of the occupations typically associated with benzene exposure.

Acetylcysteine↗
PubMed1994

[Mortality in proximal femur fractures in elderly people].

Retrospectively we studied 30 patients with fracture of the proximal femur, who were no less than 75 years old and who died during their hospitalization in the Department of Surgery of the Medical University of Lübeck from 1986 to 1991. Mortality was 6% (30/498). The mean time of death was the 13th day after administration. Nineteen patients have had 1, 7 patients 2 serious preexisting diseases. Twelve patients developed 1, 8 patients 2, and 1 patient 3 serious complications. The high amount of patients (30%, 9/30) who could not be operated reflects the great influence of the trauma itself on the mortality of the old multimorbid surgical patient. Intra-operative complications were seldom (1 pulmonary embolism), postoperative complications, however, were the limiting factors of the course of illness and are associated with a poor outcome.

Aged↗
PubMed1994

von Willebrand's disease and hemophilia are associated with diminished thromboxane A2 (TXA2) formation in clotting whole blood.

Von Willebrand's disease (vWd) and hemophilia are associated with hemorrhagic diathesis and disturbances in platelet aggregation to vessel wall. We compared the time course of thromboxane A2 (TXA2) formation by platelets during spontaneous clotting of blood of patients with von Willebrand syndrome and from patients with hemophilia A or B with that of healthy controls which were matched for sex, age and serum lipid status. In clotting blood of healthy females the TXA2 production rose at 37 degrees C in 60 min up to 228.2 +/- 32.3 ng/ml. In patients with vWd the TXA2 production at 60 min was significantly lower (129.1 +/- 26.7 ng/ml, p < 0/05). In hemophilia type A and B the TXA2 formation after 5-30 min was significantly diminished in comparison to healthy male controls (p < 0.05). From the diminished amount of TXA2 formed during spontaneous clotting of whole blood we conclude that the activation of platelets of patients with von Willebrand syndrome or hemophilia type A and B is diminished as compared to healthy controls possibly caused by reduced formation of thrombin in the blood coagulation process.

Adolescent↗
PubMed1994

[Function of the trunk musculature in elite rowers].

In sports medicine, disorders of muscle function and muscular imbalance play an important role in both risk of injury and lack of improvement. They are highly important for the trunk muscle function, since low back pain is one of the most common disorders of sportsmen and is usually diagnosed as "functional disorder" without further specification. Despite its relevance, little is known about the specific capabilities of trunk muscle function in elite sportsmen. This study describes the measurement of trunk muscle torque, velocity, endurance and coordination in elite sportsmen. The study was performed by clinical examination and an isoinertial testing device (ISOSTATION-B200, Isotechnologies, Hillsborough, NC, USA). The results of a study of 20 German elite male rowers are presented and the relevance of these results is discussed in respect of their implications for training and therapy in injured athletes. Mean torques in extension and flexion were 319 Nm and 295 Nm, respectively. Elite rowers were stronger in isometric torque in all planes compared to normals and other sports groups, such as tennis players and swimmers. The better the rowing performance was, the lower was the extension/flexion ratio, the coordination and the decrease in velocity during endurance testing. This testing procedure is a useful tool in the measurement of trunk performance in elite sportsmen.

Adult↗
PubMed1994

[Voice rehabilitation with the jejunum speech siphon: the biventer rein, a modification for prevention of aspiration].

The principle of operative voice rehabilitation after laryngectomy consists in forming a shunt between trachea and hypopharynx. The results of voice rehabilitation are generally good. Aspiration is the main disadvantage of this method, which negatively affects the quality of life. We modified the jejunum siphon of Ehrenberger and used this shunt operation mostly in total pharyngolaryngectomies. To avoid aspirations we formed a rein from both sides of the neck using the m. biventer. This rein hold the knee of the siphon and acts as a sphincter when the patient swallows, because the contractility of this muscle is preserved. We present this new method and compare the results of the first seven patients with those of the patients who were operated on without a rein from the m. biventer.

Adult↗
PubMed1994

Stimulation of a glycosyl-phosphatidylinositol-specific phospholipase by insulin and the sulfonylurea, glimepiride, in rat adipocytes depends on increased glucose transport.

Lipoprotein lipase (LPL) and glycolipid-anchored cAMP-binding ectoprotein (Gce1) are modified by glycosyl-phosphatidylinositol (GPI) in rat adipocytes, however, the linkage is potentially unstable. Incubation of the cells with either insulin (0.1-30 nM) or the sulfonylurea, glimepiride (0.5-20 microM), in the presence of glucose led to conversion of up to 35 and 20%, respectively, of the total amphiphilic LPL and Gce1 to their hydrophilic versions. Inositol-phosphate was retained in the residual protein-linked anchor structure. This suggests cleavage of the GPI anchors by an endogenous GPI-specific insulin- and glimepiride-inducible phospholipase (GPI-PL). Despite cleavage, hydrophilic LPL and Gce1 remained membrane associated and were released only if a competitor, e.g., inositol-(cyclic)monophosphate, had been added. Other constituents of the GPI anchor (glucosamine and mannose) were less efficient. This suggests peripheral interaction of lipolytically cleaved LPL and Gce1 with the adipocyte cell surface involving the terminal inositol-(cyclic)monophosphate epitope and presumably a receptor of the adipocyte plasma membrane. In rat adipocytes which were resistant toward glucose transport stimulation by insulin, the sensitivity and responsiveness of GPI-PL to stimulation by insulin was drastically reduced. In contrast, activation of both GPI-PL and glucose transport by the sulfonylurea, glimepiride, was not affected significantly. Inhibition of glucose transport or incubation of rat adipocytes in glucose-free medium completely abolished stimulation of GPI-PL by either insulin or glimepiride. The activation was partially restored by the addition of glucose or nonmetabolizable 2-deoxyglucose. These data suggest that increased glucose transport stimulates a GPI-PL in rat adipocytes.

Adipocytes↗
PubMed1994

Magnetic resonance imaging-controlled laser-induced interstitial thermotherapy.

RATIONALE AND OBJECTIVES: Laser-induced interstitial thermotherapy (LITT) may become an attractive modality for minimally invasive tumor therapy. Magnetic resonance imaging (MRI) could be used to assist this procedure. METHODS: A T1-weighted turbo fast low-angle shot (FLASH) sequence for on-line monitoring of the laser-influenced region (liver, muscle) was investigated. Sequence parameters were optimized for maximal image contrast. Magnetic resonance imaging-controlled LITT was performed in vitro, in vivo (rabbits), and in 8 human investigations (6 patients). Special laser applicators were used to establish a uniform laser light distribution. RESULTS: With the MRI sequence used, the LITT region is visualized as a bright area outlined by a dark border. This dark border corresponds to an isotherm of 45 +/- 2 degrees C depending on the sequence parameters used. CONCLUSION: With the T1-weighted turbo-FLASH sequence, MRI can be used for on-line monitoring of interstitial laser-induced thermotherapy in moving organs.

Aged↗
PubMed1994

Identification and induction of human keratinocyte-derived IL-12.

Interleukin 12 is a heterodimeric molecule that serves as a potent co-stimulator enhancing the development of Th1 cells. As one of the classical Th1 cell-mediated responses is contact sensitivity in skin, we wondered whether IL-12 might be produced by epidermal cells and serve as a mediator of this immune response. Using a sensitive, quantitative PCR technique we demonstrate that p35 chain mRNA of IL-12 is produced constitutively by human epidermal cells, whereas p40 chain mRNA can only be detected in epidermis treated with contact allergen, but not epidermis exposed to irritants or tolerogens. Time course studies showed a dramatic induction of IL-12 p40 mRNA 4 h after in vivo allergen treatment reaching peak strength after 6 h. In cell depletion assays we show that epidermal keratinocytes are the major source of this cytokine in the epidermis. This was further supported by analysis of mRNA derived from the human keratinocyte cell line HaCat expressing IL-12 p35 and p40 mRNA upon stimulation. The presence of bioactive IL-12 in supernatants derived from allergen-stimulated epidermal cells was demonstrated by IL-12-specific bioassay. Additional evidence for the functional importance of IL-12 in primary immune reactions in skin was obtained in allogeneic proliferation assays using human haptenated epidermal cells containing Langerhans cells as APC and allogeneic CD4+ T cells as responders. Anti-IL-12 mAb inhibited the proliferation of T cells by approximately 50%. In aggregate our data demonstrate that nonlymphoid keratinocytes are capable of producing functional IL-12 and provide evidence for the functional significance of IL-12 in primary immune responses in skin.

Antibodies, Monoclonal↗