PubMed Health⌕ Search

Biomedical subjects

G Krause

Publications and source records attributed to G Krause.

At least 19 recordsLinked to original sources
PubMed1995

Structure effects of double D-amino acid replacements: a nuclear magnetic resonance and circular dichroism study using amphipathic model helices.

D-Amino acid replacements and the determination of resulting structural changes are a useful tool to recognize amphipathic helices in biologically active peptides such as neuropeptide Y and corticotropin-releasing factor. In this paper the secondary structures of one amphipathic alpha-helical peptide and its double D-amino acid analog have been determined by means of 1H NMR and CD spectroscopies under equivalent conditions. The chemical shifts (NH and C alpha H) and the analysis of nuclear Overhauser effects show a split of the continuous helix for the all-L peptide into two helices at the position of double D-amino acid replacement. Hydrogen exchange rates correlate with water accessibilities in the hydrophobic/hydrophilic face and confirm the amphipathic helical structure in the all-L peptide as well as in its double D-amino acid analog. A significantly accelerated hydrogen isotope exchange rate is observed for the D-Ala9 backbone proton, implying an increased flexibility at that position. These results show that the incorporation of an adjacent pair of D-amino acids only causes a local change in structure and flexibility, which makes the double D replacement interesting as a tool for specific helix-disturbing modifications to search for helical conformations in biologically active peptides.

Amino Acid Sequence↗
PubMed1992

A Pro to His mutation in active site of thioredoxin increases its disulfide-isomerase activity 10-fold. New refolding systems for reduced or randomly oxidized ribonuclease.

Thioredoxin (Trx) from Escherichia coli was compared with bovine protein disulfide-isomerase (PDI) for its ability to catalyze native disulfide formation in either reduced or randomly oxidized (scrambled) ribonuclease A (RNase). On a molar basis, a 100-fold higher concentration of Trx than of PDI was required to give the same rate of native disulfide formation measured as recovery of RNase activity. A Pro-34 to His (P34H Trx) mutation in the active site of E. coli Trx (WCGPC), mimicking the two suggested active sites in PDI (WCGHC), increased the catalytic activity in disulfide formation about 10-fold. The mutant P34H Trx displayed a 35-mV higher redox potential (E'0) of the active site disulfide/dithiol relative to wild type Trx, making it more similar to the redox potential observed for PDI. This higher redox potential correlates well with the enhanced activity and suggests a role for the histidine side chain. Enzymatic isomerization of disulfides in scrambled, oxidized RNase requires the presence of a catalytic thiol such as GSH to initiate the thiol-disulfide interchange. Bovine thioredoxin reductase, together with NADPH, could replace GSH. For oxidative folding of reduced RNase in air with Trx, P34H Trx, or PDI, catalytic amounts of sodium selenite (1 microM) resulted in rapid disulfide formation and high yields of ribonuclease activity equivalent to previously known redox buffers of GSH and GSSG. These results demonstrate no obligatory role for glutathione in disulfide formation. A possible mechanism for the unknown thiol oxidative process accompanying folding and protein disulfide formation in vivo is discussed.

Amino Acid Sequence↗
PubMed1992

Influence of Corynebacterium parvum-PER on disease progression in the NZB/W model of systemic lupus erythematosus.

Murine models of systemic lupus erythematosus (SLE) exhibit some, but not all, of the characteristics of human disease. Disease progression in the animal models is regulated by autoantibodies, genetics and inflammatory processes. In the present study, the influence of the pyridine extract residue of Corynebacterium parvum Type I (CP-PER) on disease progression in the NZB/W model of SLE was investigated. CP-PER is known to activate the reticuloendothelial (RE) system of mice and can alter a number of host responses. Injection of NZB/W females with CP-PER at 10 weeks of age, prior to the onset of overt disease, led to a transient activation of the RE system which then appeared to return to levels observed in untreated animals. Sera from treated and untreated animals were analyzed for anti-nuclear antibodies, and anti-ssDNA antibodies. The most prominant change in the treated animals was an increase in the titer of anti-ssDNA. On average, animals in the early treatment group also lived somewhat longer than those in the untreated control group. Treatment of animals with CP-PER at 6 months of age, when the disease was evident, again led to transient RES activation and an altered autoantibody profile. Animals treated with CP-PER at 6 months of age exhibited a slightly accelerated death rate when compared with the untreated controls. These results indicate that bacterial response modifiers such as C. parvum-PER, which contains primarily the RES activating activity of C. parvum, can alter disease progression in the NZB/W model.

Animals↗
PubMed1992

Aging in humans: a continuous 20-year study of physiologic and dietary parameters.

The old adage, "You are what you eat," is not always reliable, as demonstrated in this mixed-longitudinal study of men that began in 1969. Mean values of percent body fat, total body potassium (TBK), and total serum cholesterol (SCHOL) did not show changes that correlated with any studied nutrient from repeated 4-day diet records. Mean blood pressure increased with increased body weight as age increased. High-density lipoprotein cholesterol decreased when polyunsaturated fat intake increased. The men had decreased mean height, TBK and increase in percent body fat as age increased. Food energy intake decreased equally from carbohydrates, protein and fat as age increased. Large standard deviations of mean values of measured parameters document heterogeneity of these subjects. A subset of 144 male cohorts was studied serially for 20 years in time-age, cross-sectional and longitudinal series. As they aged, height and TBK decreased (p < or = 0.05), percent body fat and blood pressure increased (p < or = 0.04) and SCHOL had no significant change. Intake of all nutrients decreased significantly (p < or = 0.03) longitudinally and with time, showing a time effect.

Adipose Tissue↗
PubMed1991

Motion of aromatic side chains, picosecond fluorescence, and internal energy transfer in Escherichia coli thioredoxin studied by site-directed mutagenesis, time-resolved fluorescence spectroscopy, and molecular dynamics simulations.

We have determined the picosecond fluorescence of the four aromatic amino acid residues (W28, W31, Y49, and Y70) in wild-type Escherichia coli thioredoxin (wt Trx) and a mutant Trx with W31 replaced by phenylalanine, Trx-W28-W31F. The internal motions of the four aromatic side chains were also analyzed. We examined the possibility of using internal energy transfer from tyrosine to tryptophan as a measure of long-range distances. The major features of the lifetime distribution of tryptophan fluorescence were unchanged in the W31F mutation, indicating that the environment of W28 is similar in both wt Trx and Trx-W28-W31F. However, the mutation of W31F changed the mobility of W28, situated close to the active-site disulfide/dithiol, but not the mobility of two tyrosines, Y49 and Y70, situated on the other side of the molecule. The mobility of the two tyrosine residues increased upon reduction of the active-site disulfide, indicating a looser structure with reduction. This increased motion could also be seen from molecular dynamics simulations. The change in energy transfer rates, as judged by tyrosine fluorescence lifetimes, was in agreement with energy transfer rates calculated from the molecular dynamics simulations. The anisotropy of tryptophan and tyrosine fluorescence could be separated in three parts: (I) overall rotation of the protein (10(-9)s), (II) internal mobility of side chains (10(-10)s), and (III) a very fast relaxation (10(-12)s). We can only experimentally detect this very fast relaxation when the internal motion is not present.

Amino Acids↗
PubMed1991

Assignment of the 15N NMR spectra of reduced and oxidized Escherichia coli thioredoxin.

As a necessary first step in the use of heteronuclear correlated spectra to obtain high resolution solution structures of the protein, assignment of the 15N NMR spectra of reduced and oxidized Escherichia coli thioredoxin (Mr 12,000) uniformly labeled with 15N has been performed. The 15N chemical shifts of backbone amide nitrogen atoms have been determined for both oxidation states of thioredoxin using 15N-1H correlated and two-dimensional heteronuclear single-quantum coherence (HSQC) TOCSY and NOESY spectra. The backbone assignments are complete, except for the proline imide nitrogen resonances and include Gly33, whose amide proton resonance is difficult to observe in homonuclear 1H spectra. The differences in the 15N chemical shift between oxidized and reduced thioredoxin, which occur mainly in the vicinity of the two active site cysteines, including residues distant in the amino acid sequence which form a hydrophobic surface close to the active site, are consistent with the differences observed for proton chemical shifts in earlier work on thioredoxin.

Amino Acid Sequence↗
PubMed1991

Mimicking the active site of protein disulfide-isomerase by substitution of proline 34 in Escherichia coli thioredoxin.

To mimic the active sites (Trp-Cys-Gly-His-Cys) contained in two thioredoxin-like domains of the eukaryotic enzyme protein disulfide-isomerase (PDI, EC 5.3.4.1), the Pro-34 residue of Escherichia coli thioredoxin (Trx) was replaced by His using site-directed mutagenesis. The mutant P34H Trx was isolated in high yield and was stable. The equilibrium between Trx and NADPH in the thioredoxin reductase (TR)-catalyzed reaction revealed that the redox potential (E'o) or P34H Trx at pH 7.0 was -235 mV as compared with -270 mV for wild type (wt) Trx. The higher E'o value made P34H Trx more similar to PDI and contributed to prominent changes in Trx functions, e.g. improved activity with TR and slower reduction of protein disulfides. Compared to wt Trx, the P34H oxidized Trx was about twice as good a substrate for TR from E. coli and four times as efficient with calf thymus TR. A novel fluorimetric assay permitted direct recording of the reaction between insulin disulfide(s) and reduced Trx. At pH 8 and 15 degrees C, second-order rate constants for wt Trx of 2 x 10(4) M-1 s-1 and for P34H Trx of 3 x 10(3) M-1 s-1 were obtained, and a different equilibrium was observed consistent with differences in E'o values. Also when the reduction mechanism of insulin was examined using NADPH and TR, P34H Trx behaved differently from wt Trx or PDI. P34H Trx may be useful as an analogue of PDI for disulfide formation in vivo and in vitro.

Amino Acid Sequence↗
PubMed1991

Substitution of the conserved tryptophan 31 in Escherichia coli thioredoxin by site-directed mutagenesis and structure-function analysis.

All prokaryotic and eukaryotic thioredoxins contain a conserved tryptophan residue, exposed at the active site disulfide/dithiol. The role of this W31 in Escherichia coli thioredoxin (Trx) was studied by site-directed mutagenesis. Four mutant Trx with W31Y, W31F, W31H, and W31A replacements were characterized. Very low tryptophan fluorescence emission from the remaining W28 was observed in all mutant Trx; reduction resulted in large, but variable increases (up to 11-fold) of fluorescence, to levels higher than in native or denatured wild-type Trx, demonstrating a previously postulated change involving W28. All W31 mutant Trx were good substrates for E. coli thioredoxin reductase. Compared with wild type, the apparent Km values were increased less than 2-fold for the W31A, W31H, and W31F Trx and the W31Y Trx showed even slightly higher catalytic efficiency (kcat/Km value). Functions of reduced Trx with ribonucleotide reductase and in reduction of insulin disulfides were more strongly influenced by the W31 replacements, in particular at low pH for A and H residues. T7 DNA polymerase activity generated by T7 gene 5 protein and reduced Trx was lowered by large factors for W31Y, W31A, or W31H compared with W31F or the wild-type protein. The in vivo function of Trx was studied by using pUC118-trxA expression in an E. coli trxA- background. The trxA genes with W31Y and W31F substitutions restored, fully and partly, the methionine sulfoxide utilization of a trxA- metE- test strain; W31A and W31H mutations resulted in no growth. Propagation of M13 was moderately impeded by W31Y and W31F or severely by W31A and W31H replacements. Growth of a phage T3/7 hybrid was possible only with the W31Y and W31F substitutions reflecting the in vitro results for T7 DNA polymerase.

Amino Acid Sequence↗
PubMed1991

Effects of cholesterol and fat modification of self-selected diets on serum lipids and their specific fatty acids in normocholesterolemic and hypercholesterolemic humans.

A 6-month crossover diet plan was employed to study the effects on human serum lipids of adding margarine or butter to otherwise self-selected diets that included two eggs daily. Two groups of subjects were studied: 51 free-living normocholesterolemic and 20 hypercholesterolemic (greater than 240 mg dl). Four-day diet records in each interval showed that subjects ate about 16% of total dietary fat as either butter or margarine. Blood samples taken every 6 weeks showed variable mean serum total cholesterol (SCHOL), high-density-lipoprotein cholesterol (HDL-C), and serum triglycerides (STG). The normocholesterolemic subjects who ate butter first had by 24 weeks mean SCHOL values equal to their entry values; those who ate margarine first had increased SCHOL values throughout the study. By the end of the study, the hypercholesterolemic subjects showed either no change or a slight decrease in both SCHOL and HDL-C values. Specific fatty acids were distributed differently in the serum fractions of triacylglycerol (TGFA), cholesteryl esters (CEFA), and phospholipids (PLFA). These distributions remained constant in both normocholesterolemic and hypercholesterolemic subjects regardless of the type and amount of fat consumed.

Adult↗
PubMed1991

Influence of biological response modifiers of bacterial origin on disease progression in the MRL-lpr model of systemic lupus erythematosus.

Murine models of systemic lupus erythematosus exhibit some, but not all of the characteristics of human disease. Disease progression in the animal models is related to autoantibodies, genetics, and inflammatory processes. In this report the effects of two bacterial biological response modifiers (BRM) on disease progression in the MRL-lpr model were investigated. The two BRM tested were C. parvum and Bacillus-Calmette-Guerin (BCG), both of which are stimulators of the reticuloendothelial system and both of which have been shown by others to influence disease progression in NZB/W mice. Treatment of 10-week-old mice with C. parvum led to transient alterations in hepatosplenomegaly and plasma proteinase regulation, which then returned to control values. Treatment with BCG led to even more transient effects on the mice. Neither BRM appeared to impact on disease-associated alterations in autoantibody titres, hepatosplenomegaly, or elevations in plasma proteinase activity. Likewise, treatment of 17-week-old MRL-lpr mice with C. parvum did not influence disease progression as evidenced by survival, autoantibody production, or hepatosplenomegaly. Therefore, in contrast to the NZB/W strain, treatment of the MRL-lpr strain with these BRM does not appear to impact on disease progression. This difference may be due to the influence of the lpr accelerator gene in this model.

Aging↗
PubMed1989

Total body potassium in aging humans: a longitudinal study.

Total body potassium (TBK) data calculated from longitudinal measurements over 18 y of 40K by whole-body counting of 564 male and 61 female healthy humans in a 2-pi liquid scintillation counter show little change in females younger than 50 y compared with males of those ages. Males show less TBK from 41 y onward as they age, with most rapid rate of loss between 41 and 60 y. Females have a rapid loss of TBK when they are older than 60 y; the loss is at a greater rate than that of males. Percent total body fat calculated from total body weight and lean body mass (LBM) derived from TBK document greater adiposity in females at all ages except ages 51-60 y when females are similar to males in change in percent fat per year per centimeter.

Adipose Tissue↗
PubMed1989

[Leakage measurements with the dye extraction method and direct methotrexate level determination (enzyme immunoassay) during and after regional cytostatic limb perfusion].

The dye stuff extinction-method stood the test for checking the separation between the regional and the systemic circulation during the perfusion of extremity tumors by cytostatic drugs. On the other hand the use of an enzyme immunoassay for direct methotrexate-level measuring in serum is recommendable in the postoperative phase. Complications by methotrexate are recognized early by means of an enzyme immunoassay and an individualized application is possible.

Chemotherapy, Cancer, Regional Perfusion↗
PubMed1988

Effect of melatonin on intraocular pressure.

We studied the effect of orally administered melatonin on intraocular pressure in humans. We suppressed serum melatonin levels by exposing our subjects to bright light. Our experiments suggest that melatonin lowers intraocular pressure in man. This may prove to be a therapeutically useful agent since melatonin appears to be relatively free of side effects and is effective in small quantities.

Administration, Oral↗
PubMed1987

Cerebral endothelial microvilli following global brain ischemia in dogs.

Cerebral blood vessels (BVs) of dogs subjected to global brain ischemia by complete cardiac arrest of 15 min followed by 8 h of reperfusion, were studied in neocortex and hippocampus by means of transmission electron microscopy. Widespread endothelial microvilli were present in the postischemic animals. The number of endothelial microvilli in the postischemic animals (mean/BV in the neocortex = 3.26 and in the hippocampus = 2.54) was significantly larger than that in the non-ischemic controls (mean/BV in the neocortex = 1.39 and in the hippocampus = 0.84), P for both regions being less than 0.05. Arterioles, venules and capillaries, all were equally affected. Endothelial pinocytotic vesicles were also observed frequently in the postischemic dogs. Marked pericapillary swelling of astrocytic foot processes was present in the surrounding neuropil. It is concluded that the prominent cerebral endothelial microvilli recognized after 8 h of reperfusion following cardiac arrest in this experimental model of global brain ischemia, may play a significant role in the development of delayed postischemic hypoperfusion.

Animals↗
PubMed1987

Effect of flunarizine on global brain ischemia in the dog: a quantitative morphologic assessment.

The effects of flunarizine, a calcium antagonist, were evaluated in an experimental model of global brain ischemia produced by 15 min of cardiac arrest followed by resuscitation and reperfusion. One group of dogs received flunarizine (0.1 mg/kg intravenously during a 10-min period) at the onset of resuscitation. Another group of dogs underwent cardiac arrest, resuscitation, and reperfusion but did not receive flunarizine. A third group served as nonischemic control. In situ-fixed brains of all animals (nonischemic controls and the postischemic dogs after 8 h of reperfusion) were examined for anoxic ischemic injury. Quantitation of the ischemic neurons was carried out in parietal cortex, hippocampus, and cerebellum by using an image analysis system. Significant difference in the number of necrotic neurons between the flunarizine-treated group and the ischemic controls was noted in the hippocampus only; the mean percentage of necrotic neurons in the two groups being 14.8 +/- 9.6 and 29.3 +/- 12.1, respectively (P less than 0.05). These results indicate that flunarizine has an ameliorating effect on neuronal injury in the hippocampus that follows cardiac arrest in this experimental model of global brain ischemia. However, flunarizine was not found to be effective in reducing the ischemic neuronal damage in the cortex or the cerebellum.

Animals↗
PubMed1986

Heroin.

Explore the source record for details and available documents.

Attitude of Health Personnel↗