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Biomedical subjects

C Wolff

Publications and source records attributed to C Wolff.

At least 109 records · Page 6Linked to original sources
PubMed1989

Fluorescence energy transfer detects changes in fibronectin structure upon surface binding.

We report here the changes in intramolecular distances in human plasma fibronectin (Fn) detected, upon adsorption of the protein to the surface of the Cytodex dextran microcarrier, using a fluorescence energy transfer technique. The glutamine-3 residue, near the amino terminus of each chain, was labeled enzymatically with either monodansylcadaverine (dansyl) or monofluoresceinyl-cadaverine (fluorescein) by use of coagulation factor XIIIa. Using this donor (dansyl)-acceptor (fluorescein) pair, and steady-state measurements, we demonstrated previously that the two amino termini of plasma fibronectin in solution were juxtaposed and separated by 23 A (C. Wolff and C.-S. Lai (1988) Biochemistry 27, 3483-3487). Upon adsorption to the microcarrier, the energy transfer was found to be completely abolished, suggesting that the surface binding induces a conformational change by which the distance between the two amino termini is increased to more than 70 A. Moreover, we have labeled the amino terminus of each chain with fluorescein and the two free sulfhydryl groups of each chain with coumarinyl-phenylmaleimide which serves as an energy donor. The emission spectra of the double-labeled protein in solution showed the occurrence of energy transfer, indicating that the relative distances between the amino termini and the free sulfhydryl group(s) are within 70 A. Upon surface binding, a decrease in the energy transfer between this donor-acceptor pair was also noted. The results presented here are consistent with the notion that plasma Fn undergoes a drastic conformational change upon surface binding, perhaps changing from a compact form to an extended form. This process may be important for the surface activation of the fibronectin molecule.

Energy Transfer↗
PubMed1988

Evidence that the two amino termini of plasma fibronectin are in close proximity: a fluorescence energy transfer study.

A fluorescence energy transfer technique has been used to study the intramolecular distance between the two amino termini of human plasma fibronectin. The glutamine-3 residue near the amino terminus of each chain was labeled enzymatically with either monodansylcadaverine or monofluoresceinylcadaverine by use of coagulation factor XIIIa. The nonradiative fluorescence energy transfer between the dansyl (donor) and fluorescein (acceptor) pair in the same protein molecule was determined from steady-state fluorescence measurements. On the basis of the transfer efficiency of 78%, the intramolecular distance between two glutamine-3 residues of fibronectin was estimated to be approximately 23 A, suggesting that the two amino termini of plasma fibronectin are in close proximity. High salt, which affects the hydrodynamic properties of the protein, has no effect on the measured distance. The results support the contention that both compact (in low salt) and expanded (in high salt) conformers of fibronectin are relatively spherical in shape.

Energy Transfer↗
PubMed1988

Serum proline and blood lactate levels in alcoholic patients without hepatic failure: relationship with alcohol ingestion and histological activity.

It has not yet been established whether serum proline and blood lactate levels are increased in alcoholic liver disease. We measured serum proline and blood lactate in controls and in patients with different stages of alcoholic liver disease in the absence of hepatic failure. Samplings were done in both abstinent and drinking alcoholics. Compared to controls, there was a striking increase in serum proline levels in 52 abstinent alcoholics with little or no hepatic fibrosis by histological assessment (0.10 +/- 0.01 vs. 0.155 +/- 0.008; p less than 0.005). Blood lactate levels were within the normal range and did not correlate with serum proline levels. On the other hand, serum proline and blood lactate levels were independent of hepatic necrosis and inflammation scores. In addition, in 10 patients with blood alcohol concentrations between 0.3 mg/ml and 7.8 mg/ml, serum lactate and proline were significantly elevated (2.42 +/- 0.29 mg/ml and 0.275 +/- 0.0026 mg/ml, respectively; p less than 0.005). These results show that there is an association between serum proline levels and the abstinence period in alcoholic patients. They further suggest that in alcoholic patients neither serum proline nor blood lactate concentrations are reliable markers for liver histological activity (necrosis and inflammation) or fibrosis.

Adult↗
PubMed1988

Aspartate taxis mutants of the Escherichia coli tar chemoreceptor.

The Tar protein of Escherichia coli belongs to a family of methyl-accepting inner membrane proteins that mediate chemotactic responses to a variety of compounds. These transmembrane signalers monitor the chemical environment by means of specific ligand-binding sites arrayed on the periplasmic side of the membrane, and in turn control cytoplasmic signals that modulate the flagellar rotational machinery. The periplasmic receptor domain of Tar senses two quite different chemoeffectors, aspartate and maltose. Aspartate is detected through direct binding to Tar molecules, whereas maltose is detected indirectly when complexed with the periplasmic maltose-binding protein. Saturating levels of either aspartate or maltose do not block behavioral responses to the other compound, indicating that the detection sites for these two attractants are not identical. We initiated structure-function studies of these chemoreceptor sites by isolating tar mutants which eliminate aspartate or maltose taxis, while retaining the ability to respond to the other chemoeffector. Mutants with greatly reduced aspartate taxis are described and characterized in this report. When present in single copy in the chromosome, these tar mutations generally eliminated chemotactic responses to aspartate and structurally related compounds, such as glutamate and methionine. Residual responses to these compounds were shifted to higher concentrations, indicating a reduced affinity of the aspartate-binding site in the mutant receptors. Maltose responses in the mutants ranged from 10 to 80% of normal, but had no detectable threshold shifts, indicating that these receptor alterations may have little effect on maltose detection sensitivity. The mutational changes in 17 mutants were determined by DNA sequence analysis. Each mutant exhibited a single amino acid replacement at residue 64, 69, or 73 in the Tar molecule. The wild-type Tar transducer contains arginines at all three of these positions, implying that electrostatic forces may play an important role in aspartate detection.

Aspartic Acid↗
PubMed1988

Maltose chemoreceptor of Escherichia coli: interaction of maltose-binding protein and the tar signal transducer.

The maltose chemoreceptor in Escherichia coli consists of the periplasmic maltose-binding protein (MBP) and the Tar signal transducer, which is localized in the cytoplasmic membrane. We previously isolated strains containing malE mutations that cause specific defects in the chemotactic function of MBP. Four of these mutations have now been characterized by DNA sequence analysis. Two of them replace threonine at residue 53 of MBP with isoleucine (MBP-TI53), one replaces an aspartate at residue 55 with asparagine (MBP-DN55), and the fourth replaces threonine at residue 345 with isoleucine (MBP-TI345). The chemotactic defects of MBP-TI53 and MBP-DN55, but not of MBP-TI345, are suppressed by mutations in the tar gene. Of the tar mutations, the most effective suppressor (isolated independently three times) replaces Arg-73 of Tar with tryptophan. Two other tar mutations that disrupt the aspartate chemoreceptor function of Tar also suppress the maltose taxis defects associated with MBP-TI53 and MBP-DN55. One of these mutations introduces glutamine at residue 73 of Tar, the other replaces arginine at residue 69 of Tar with cysteine. These results suggest that regions of MBP that include residues 53 to 55 and residue 345 are important for the interaction with Tar. In turn, arginines at residues 69 and 73 of Tar must be involved in the recognition of maltose-bound MBP and/or in the production of the attractant signal generated by Tar in response to maltose-bound MBP.

ATP-Binding Cassette Transporters↗
PubMed1988

Increased lipid peroxidation in hyperthyroid patients: suppression by propylthiouracil treatment.

Plasma and urinary levels of thiobarbituric acid reactive substances (TBAR) were determined in 24 hyperthyroid patients, 19 hypothyroid subjects, 35 controls, and 17 hyperthyroid patients before and after propylthiouracil (PTU) treatment (400 mg/day for 2-3 months), as indexes of lipid peroxidation. These measurements were carried out together with t-butyl hydroperoxide (t-BHP)-induced oxygen uptake and visible chemiluminescence in erythrocytes as functional tests related to the antioxigenic capacity of cells. Hyperthyroid patients exhibited increased levels of plasma and urinary TBAR compared to controls. Erythrocyte suspensions from hyperthyroid patients showed, compared to controls, higher rates of oxygen consumption with shorter induction periods upon addition of t-BHP, together with 142% and 75% increases in basal and t-BHP-induced chemiluminescence, respectively. Levels of TBAR in untreated hyperthyroid patients in plasma (16.2 +/- 1.3 pmol/mg of protein) and urine (15.9 +/- 1.5 nmol/mg of creatinine) were decreased after PTU treatment (Plasma, 9.5 +/- 0.7, p less than 10(-4); urine, 7.8 +/- 0.9, P less than 10(-5) to values not significantly different from those of the control group (plasma, 10.3 +/- 0.6; urine, 7.9 +/- 0.7). Compared to control, elevated rates of oxygen uptake induced by t-BHP, basal and t-BHP-induced chemiluminescence in erythrocyte suspensions from untreated hyperthyroid patients were reverted to normal by PTU, while decreased induction period (T0) values were enhanced. Determination of these lipid peroxidative parameters in hypothyroid patients revealed no significant changes over control values, excepted t-BHP-induced chemiluminescence in erythrocytes that was diminished. These data indicate that hyperthyroidism is associated with a pro-oxidant condition characterized by an enhancement in circulating and urinary lipid peroxidative indexes, which is suppressed by PTU treatment. It is suggested that this condition might reflect an oxidative stress at cellular level in tissues which are target for thyroid hormone action with a calorigenic response.

Adult↗
PubMed1987

Relationship between hepatic levels of glutathione and sulphobromophthalein retention in hyperthyroidism.

1. Sulphobromophthalein (BSP) retention and liver glutathione levels were studied in uncomplicated hyperthyroid patients. 2. BSP retention was increased in 52.5% of the subjects at admission (n = 40) and in 28% of the cases after 3 months of propylthiouracil treatment (300-400 mg/day) (n = 25). 3. Hepatic levels of glutathione were measured in six patients and significant inversed power correlation with BSP retention was observed (r = 0.968, P less than 0.001). 4. These data support the contention that the alteration of the BSP retention observed in hyperthyroidism could be due to a decreased availability of hepatic glutathione for conjugation.

Adolescent↗
PubMed1987

[The fate of patients with diabetes in pregnancy--classification of gestational diabetes following the completion of pregnancy (p.g. classification)].

In 69 patients with a gestational diabetes-diagnosed by reproducible pathological results of two oral glucose loads (50 g) or by fasting blood glucose values of greater than or equal to 6.7 mmol/l in pregnancy the carbohydrate metabolism was checked postgestationally again with a glucose tolerance test (75 g) in a period of 6 weeks up to 2 years post partum. The postgestational classification showed the following results: manifested diabetes n = 15 (21.7%), impaired glucose tolerance n = 15 (21.7%), non-classificable disturbed carbohydrate tolerance n = 10 (14.5%), normal test results n = 29 (42%). The high rate of diabetic manifestations underlines the necessity of a postgestational classification of the so-called gestational diabetes controlled by the delivering center. The high risk of manifestation is calculable in the whole group, but not predictable for the single case. The risk is the higher the earlier a glucosuria in pregnancy can be found (before the 24th week), the earlier an insulinisation is necessary to guarantee a normoglycemia and the higher individual deviations of the individual blood glucose values during the daily course are observed (measurable with the glycemic index acc. Michaelis et al.). Additional risk factors are: obesity, an age over 30 years at the beginning of pregnancy, and heredity of first degree. From the retrospective point of view of a postgestational classification new therapeutic aspects could not be verified to avoid diabetes manifestation. Nevertheless an exact normoglycemic control and a very early start of treatment, a correct screening of risk factors and an immediate diagnosis of a gestational diabetes are a supposition to avert hyperglycemic dangers from the child.

Adult↗
PubMed1985

[Care of pregnant diabetic patients--results and problems].

This is a report on results and problems in taking care of pregnant diabetics in the delivering center Leipzig with special regard of women with a gestational diabetes. In the period between 1971 and 1983 a total of 314 patients were treated with manifest diabetes or a gestational diabetes. We investigated in with manifest diabetes or a gestational diabetes. We investigated in non-selected cases the morbidity of children and the perinatal mortality. The evaluation took into consideration among others the quality of metabolic guidance, the possible causes of perinatal mortality and the therapeutical indications in gestational diabetes. Depended on the changed concept of the care system we considered separately the results of two periods (first period: 1971 until 1978, 136 deliveries; second period: 1979 until 1983, 178 deliveries). We could demonstrate that a complex interdisciplinary care system can reduce the perinatal mortality and morbidity. But it is necessary to diagnose patients with a gestational diabetes earlier than actually usual.

Acidosis↗
PubMed1984

The influence of sequences adjacent to the recognition site on the cleavage of oligodeoxynucleotides by the EcoRI endonuclease.

We have investigated the influence of the nucleotide sequence adjacent to the recognition site on the rate of cleavage of DNA by the restriction endonuclease EcoRI. For this purpose two decadeoxynucleotides, d(G-G-G-A-A-T-T-C-T-T) (Ia) and d(A-A-G-A-A-T-T-C-C-C) (Ib) were synthesized. The duplex Ia X Ib is cleaved by EcoRI preferentially in the dA-rich strand (approximately 10 times over the dG-rich strand). The individual nucleotides Ia and Ib are also cleaved by EcoRI, Ib at a higher rate than Ia and both at a lower rate than Ia X Ib. The temperature dependence of the reaction rate shows that only double-stranded oligodeoxynucleotides are substrates for the EcoRI endonuclease. We have, furthermore, synthesized oligomers of d(G-G-A-A-T-T-C-C), which contain two, three and four EcoRI sites, respectively. These oligodeoxynucleotides are preferentially cleaved at the sites next to the 5' end, where the recognition site is only flanked by one dG X dC base pair, in contrast to the other sites which are flanked by three such pairs. These data indicate that sequences adjacent to the recognition site influence the rate of cleavage: dA X dT base pairs enhance and dG X dC base pairs slow down the hydrolytic activity of the EcoRI endonuclease.

Base Composition↗