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Biomedical subjects

G Fischer

Publications and source records attributed to G Fischer.

At least 217 records · Page 12Linked to original sources
PubMed1997

Probing substrate backbone function in prolyl oligopeptidase catalysis--large positional effects of peptide bond monothioxylation.

Site-specific effects on the catalytic activity of prolyl oligopeptidase from human placenta were studied using oligopeptide substrates in which a peptide bond has been replaced by a thioxo peptide bond. Two series of tetrapeptide-4-nitroanilides, Ala-Gly-Pro-Phe-NH-Np and Ala-Ala-Pro-Phe-NH-Np, along with all possible monothioxylated derivatives, were synthesised and k(cat) and Km values were determined for proteolytic cleavage at the Pro-Phe bond. Regardless of either Gly or Ala in the P2 subsite, tetrapeptides were rendered uncleavable by thioxylation at the Pro-Phe linkage. As a result, Ala-Xaa-Pro-psi[CS-NH]-Phe-NH-Np (Xaa = Gly or Ala) displayed competitive inhibition with Ki-values of 12 microM and 44 microM, respectively. Furthermore, in controlling proteolytic susceptibility of the substrates, cooperation of the P3-P2 thioxylation site and the side chain at the P2 subsite was obtained. Thioxylation at this position enhanced k(cat)/Km fivefold in the Gly series, but led to a 1.7-fold decrease in the Ala series of substrates. With respect to the Xaa-Pro peptide bond, all of the substrates underwent cis/trans isomerisation, thus presenting two stable conformers to the protease. However, the magnitudes of the isomerisation constants suggested that neither isomerisation rates nor cis/trans equilibria can explain the effect of thioxylation on the steady-state constants of proteolysis.

Catalysis↗
PubMed1997

[Neurosarcoma associated with neurofibromatosis 1. Apropos of a case and review of the literature].

BACKGROUND: Type 1 neurofibromatosis considerably increases the risk of cancer development, particularly neurosarcoma. We report a case in a patient with chemosensitive metastatic neurosarcoma. CASE REPORT: A young female patient with familial type 1 neurofibromatosis developed pleural metastasis of a neurosarcoma located on the arm. This tumor was initially highly sensitive to chemotherapy, but relapse occurred. DISCUSSION: Follow-up in the order members of the family was particularly difficult to organize. One sister developed cerebral astrocytoma. Neurosarcomas develop earlier in patients with type 1 neurofibromatosis, worsening prognosis. We suggest a prospective and structured registration of such cases using a network of clinicians and pathologists in order to improve management schemes.

Adult↗
PubMed1997

Cooperative and competitive interactions of regulatory elements are involved in the control of divergent transcription of human Col4A1 and Col4A2 genes.

The genes COL4A1 and COL4A2, coding for the two subunit chains alpha1(IV) and alpha2(IV) of collagen IV [alpha1(IV)2alpha2(IV)] are found closely linked on the human chromosome 13 in a unique head-to-head arrangement resulting in opposite strand transcription starting from a shared promoter region. Transient transfection experiments defined a shared promoter and two symmetrically arranged, downstream located and gene-specific activating elements in each gene. The shared promoter does not exhibit any transcriptional activity and efficient transcription depends on the cooperative effect of downstream elements. Mutual inhibitory effects between the two activating elements indicate competitive interactions with the shared promoter. Symmetry, cooperativity and competitivity of cis-elements are also reflected by the binding of transacting factors to the promoter and activating elements. From these data we propose a model for the coordination of divergent transcription of COL4 genes based on the cooperative and competitive interactions of the shared promoter and gene-specific regulating elements.

Binding, Competitive↗
PubMed1997

Cyclophilin A complexed with a fragment of HIV-1 gag protein: insights into HIV-1 infectious activity.

BACKGROUND: Cyclophilin A (CyPA), a receptor of the immunosuppressive drug cyclosporin A, catalyzes the cis-trans isomerization of peptidyl-prolyl bonds and is required for the infectious activity of human immunodeficiency virus type 1 (HIV-1). The crystal structure of CyPA complexed with a fragment of the HIV-1 gag protein should provide insights into the nature of CyPA-gag interactions and may suggest a role for CyPA in HIV-1 infectious activity. RESULTS: The crystal structure of CyPA complexed with a 25 amino acid peptide of HIV-1 gag capsid protein (25-mer) was determined and refined to an R factor of 0.195 at 1.8 A resolution. The sequence Ala88-Gly89-Pro90-Ile91 of the gag fragment is the major portion to bind to the active site of CyPA. Two residues of the 25-mer (Pro90-Ile91) bind to CyPA in a similar manner to two residues (Pro-Phe) of the CyPA substrate, succinyl-Ala-Ala-Pro-Phe-p-nitroanilide (AAPF). However, the N-terminus of the 25-mer (Ala88-Gly89) exhibits a different hydrogen-bonding pattern and molecular conformation than AAPF. The peptidyl-prolyl bond between Gly89 and Pro90 of the 25-mer has a trans conformation, in contrast to the cis conformation observed in other known CyPA-peptide complexes. The residue preceding proline, Gly89, has an unfavorable backbone conformation usually only adopted by glycine. CONCLUSIONS: The unfavorable backbone conformation of Gly89 of the gag 25-mer fragment suggests that binding between HIV-1 gag protein and CyPA requires a special sequence, Gly-Pro. Thus, in HIV-1 infectivity, CyPA is likely to function as a chaperone, rather than as a cis-trans isomerase. However, the observation of similarities between the C termini of the 25-mer and the substrate AAPF means that the involvement of the cis-trans isomerase activity of CyPA cannot be completely ruled out.

Amino Acid Isomerases↗
PubMed1997

Cooperation of enzymatic and chaperone functions of trigger factor in the catalysis of protein folding.

The trigger factor of Escherichia coli is a prolyl isomerase and accelerates proline-limited steps in protein folding with a very high efficiency. It associates with nascent polypeptide chains at the ribosome and is thought to catalyse the folding of newly synthesized proteins. In its enzymatic mechanism the trigger factor follows the Michaelis-Menten equation. The unusually high folding activity of the trigger factor originates from its tight binding to the folding protein substrate, as reflected in the low Km value of 0.7 microM. In contrast, the catalytic constant kcat is small and shows a value of 1.3 s(-1) at 15 degrees C. An unfolded protein inhibits the trigger factor in a competitive fashion. The isolated catalytic domain of the trigger factor retains the full prolyl isomerase activity towards short peptides, but in a protein folding reaction its activity is 800-fold reduced and no longer inhibited by an unfolded protein. Unlike the prolyl isomerase site, the polypeptide binding site obviously extends beyond the FKBP domain. Together, this suggests that the good substrate binding, i.e. the chaperone property, of the intact trigger factor is responsible for its high efficiency as a catalyst of proline-limited protein folding.

Amino Acid Isomerases↗
PubMed1997

Genetic instability and its possible evolutionary implications on the chromosomal structure of Streptomyces.

Recent progress in the understanding of the chromosomal rearrangements in Streptomyces has allowed us to envisage the possible involvement of genetic instability in the evolution of the chromosomal structure. The characterization of recombinational events in the terminal parts of the S ambofaciens chromosome, as well as the observation by other groups that linear plasmids and chromosomes are able to interact, would provide an explanation for the very high levels of polymorphism seen in the terminal inverted repeats of different strains of Streptomyces.

Chromosomes, Bacterial↗
PubMed1997

Modulation of cyclosporin A/cyclophilin interactions by drug vehicles.

Cyclosporin A (CsA) is a tight-binding inhibitor of the peptidyl-prolyl cis/trans isomerase (PPIase) activity of human cytosolic cyclophilin (Cyp18), the putative receptor for immunosuppressive effects of the drug. We examined the influence of cremophor EL (CEL), a surfactant that has found wide use for CsA formulation, on the kinetics of inhibition of the enzyme by CsA. Stock solutions of CsA in CEL administered into aqueous PPIase assays led to inhibition kinetics reminiscent to those of CsA dissolved in tetrahydrofurane, but caused an increase in the final Ki value of about sevenfold at 0.33% (v/v) CEL. The diminished drug affinity to Cyp18 obtained in experiments using CEL could also be established for analogues of cyclosporin A such as [Ala2]-Cs,[Thr2]-Cs, and [MeAla6]-Cs, exhibiting Ki values 13-16-fold higher than in the absence of CEL. In addition, the time-dependent pattern of inhibition indicate only a minor population of bioactive conformation of CsA in bulky CEL. Conformational reshuffling of the bioinactive [cis-MeLeu9-MeLeu10]-Cs to create an inhibitory fraction of the drug was delayed in the presence of CEL micelles, despite potential ability of micelles exists to catalyze cis/trans isomerizations of N-alkyl peptide bonds. The pattern of inhibition when using cyclophilins distinct in their amino acid sequences to the human enzyme can be rationalized in terms of exceptional high structural requirements of human Cyp18 for the drug conformation.

Amino Acid Isomerases↗
PubMed1997

Excellent long-term survival after allogeneic marrow transplantation in patients with severe aplastic anemia.

Between 1982 and 1996, 20 patients (10 male, 10 female) with severe aplastic anemia (SAA) with a median age of 25 years (17-37 years), received grafts from an HLA-identical sibling (n = 17), HLA-identical unrelated donor (n = 2) or identical twin (n = 1). The median time from diagnosis to marrow transplantation (BMT) was 15 months (range 1-96 months). More than half of the patients had received more than 10 units of red blood cells or platelet transfusions prior to BMT. Pretransplant immunosuppression consisted of cyclophosphamide (CY) alone (n = 10), CY in combination with total body irradiation (n = 8), and CY and antithymocyte globulin (n = 2). For graft-versus-host disease (GVHD) prophylaxis methotrexate (MTX) alone (n = 9) or MTX with cyclosporin A (n = 10) were given. One patient died on day 18 after marrow grafting due to infection; all other patients had complete and sustained engraftment (95%). Eight patients developed acute GVHD (42%), nine patients chronic GVHD (53%) including four with extensive disease manifestation. One patient experienced a secondary malignancy 11 years after BMT. Eighteen patients followed for a median of 9.45 years (0.42-14.7 years) have sustained hematological reconstitution and are alive and well with a Karnofsky performance score of at least 90%. Thus, excellent long-term survival and low morbidity make allogeneic or syngeneic BMT the treatment of choice for younger patients with severe aplastic anemia.

Adolescent↗
PubMed1997

Effect of calcitriol in combination with corticosterone, interleukin-1beta, and transforming growth factor-beta1 on nerve growth factor secretion in an astroglial cell line.

In astrocytes, nerve growth factor (NGF) synthesis has been described to be stimulated by the cytokines interleukin-1beta (IL-1beta) and transforming growth factor-beta1 (TGF-beta1) and inhibited by corticosterone. As all three factors are present in the brain under certain conditions, we investigated the effect of their combined application on NGF secretion in the astroglial cell line RC7 and, in addition, studied the effect of calcitriol (1alpha,25-dihydroxyvitamin D3). Calcitriol stimulated NGF secretion, whereas corticosterone reduced basal levels of NGF secretion as well as inhibited the NGF secretion induced by IL-1beta, calcitriol, and TGF-beta1. Calcitriol had an additive effect when applied together with IL-1beta and a synergistic effect when applied with TGF-beta1. Moreover, calcitriol not only counteracted the inhibitory effect of corticosterone on NGF secretion stimulated by TGF-beta1 but even augmented it to a level more than threefold higher than that reached with TGF-beta1 alone. Due to the trophic effect of NGF on basal forebrain cholinergic neurons, these findings might be of therapeutic relevance under conditions where cholinergic function is impaired and the endogenous levels of corticosterone, IL-1beta, or TGF-beta1 are elevated.

Animals↗
PubMed1997

Effects of a dobutamine-induced increase in splanchnic blood flow on hepatic metabolic activity in patients with septic shock.

BACKGROUND: Septic shock leads to increased splanchnic blood flow (Qspl) and oxygen consumption (VO2spl). The increased Qspl, however may not match the splanchnic oxygen demand, resulting in hepatic dysfunction. This concept of ongoing tissue hypoxia that can be relieved by increasing splanchnic oxygen delivery (DO2spl), however, was challenged because most of the elevated VO2spl was attributed to increased hepatic glucose production (HGP) resulting from increased substrate delivery. Therefore the authors tested the hypothesis that a dobutamine-induced increase in Qspl and DO2spl leads to increased VO2spl associated with accelerated HGP in patients with septic shock. METHODS: Twelve patients with hyperdynamic septic shock in whom blood pressure had been stabilized (mean arterial pressure > or = 70 mmHg) with volume resuscitation and norepinephrine received dobutamine to obtain a 20% increase in cardiac index (CI). Qspl, DO2spl, and VO2spl were assessed using the steady-state indocyanine green clearance technique with correction for hepatic dye extraction, and HGP was determined from the plasma appearance rate of stable, non-radio-active-labeled glucose using a primed-constant infusion approach. RESULTS: Although the increase in CI resulted in a similar increase in Qspl (from 0.91 +/- 0.21 to 1.21 +/- 0.34l.min-1.m2; P < 0.001) producing a parallel increase of DO2spl (from 141 +/- 33 to 182 +/- 44 ml.min-1.m2; P < 0.001), there was no effect on VO2spl (73 +/- 16 and 82 +/- 21 ml.min-1.m2, respectively). Hepatic glucose production decreased from 5.1 +/- 1.6 to 3.6 +/- 0.9 mg.kg-1.min-1 (P < 0.001). CONCLUSIONS: In the patients with septic shock in whom blood pressure had been stabilized with volume resuscitation and norepinephrine, no delivery-dependency of VO2spl could be detected. Oxygen consumption was not related to the accelerated HGP either, and thus the concept that HGP dominates VO2spl must be questioned in well-resuscitated patients with septic shock.

Adrenergic beta-Agonists↗
PubMed1997

Occurrence of deletions, associated with genetic instability in Streptomyces ambofaciens, is independent of the linearity of the chromosomal DNA.

The chromosomal structures of mutant strains of Streptomyces ambofaciens which have arisen from genetic instability were investigated by using pulsed-field gel electrophoresis and probing with sequences cloned from the unstable region which maps near the ends of the linear chromosomal DNA. The chromosomal structures of seven mutant strains harboring large deletions were classified into three types. (i) Deletions internal to one chromosomal arm were characterized in two of the mutant strains. In these strains, a linear chromosomal structure was retained, as were parts of the terminal inverted repeats sequences (TIRs) and the proteins bound to them. (ii) Four of the mutants presented a deletion including all sequences from the TIRs. A junction fragment homologous to sequences originating from internal region of both arms was characterized. Consequently, the chromosomal DNA of these strains was deduced to be circularized. Furthermore, chromosomal stability was assessed in the progeny of these circular DNA mutants. Additional deletion events were detected in 11 mutants among the 13 strains isolated, demonstrating that circular chromosomes do not correspond to a stabilization of the chromosome structure and that the occurrence of deletion could be independent of the presence of chromosomal ends. (iii) A mutant with DNA amplification was shown to have a linear chromosome with a deletion of all sequences between the amplified region and the end of the chromosome. The other chromosomal arm remained unaffected by deletion and associated with protein.

Bacterial Proteins↗
PubMed1997

Brain glucose metabolism with [18F]-fluorodeoxyglucose and positron emission tomography before and after surgical resection of epileptogenic cavernous angiomas.

Surgery of cavernous angiomas is often proposed due to the epileptogenic and hemorrhagic potential of this lesion. Little information is available on the impact of surgical resection on brain metabolism, locally or at a distance from the lesion. Fifteen patients presenting with a cavernoma and epileptic seizures underwent positron emission tomography examinations before and 1 year after surgical resection. We studied the quantitative cerebral metabolic rate of glucose (CMRGlu) with [18F]-fluorodeoxyglucose. The global brain CMRGlu remained unchanged after surgery. There was a significant decrease of metabolism in the lesion hemisphere, which remained unchanged after surgery. The perilesional regions were less metabolic than the contralateral ones and less metabolic than remote regions within the same hemisphere, before and after surgery. The absence of pre- to postsurgical variations suggests that the metabolic consequences of the lesion are maintained despite the surgical procedure during long-term follow-up studies.

Brain↗
PubMed1997

Lipohexin, a new inhibitor of prolyl endopeptidase from Moeszia lindtneri (HKI-0054) and Paecilomyces sp. (HKI-0055; HKI-0096). I. Screening, isolation and structure elucidation.

Lipohexin was isolated as a novel lipohexapeptide (I) (C39H68N6O9) from three fungal strains, Moeszia lindtneri HKI-0054, Paecilomyces sp. HKI-0055 and Paecilomyces sp. HKI-0096. The structure was elucidated by detailed mass spectrometric and NMR experiments. The proline-containing peptide displays moderate antibacterial activity against Bacillus subtilis ATCC 6633 and inhibits competitively the prolyl endopeptidase from human placenta.

Anti-Bacterial Agents↗
PubMed1997

Lipohexin, a new inhibitor of prolyl endopeptidase from Moeszia lindtneri (HKI-0054) and Paecilomyces sp. (HKI-0055; HKI-0096). II. Inhibitory activities and specificity.

The new proline-containing lipohexapeptide lipohexin (I) isolated from three fungal strains, Moeszia lindtneri (HKI-0054) and Paecilomyces sp. (HKI-0055 and HKI-0096) is a competitive inhibitor of prolyl endopeptidase (PEP) from human placenta with IC50 of 3.5 microM. Specificity of lipohexin (I) is indicated by the much weaker inhibitory activity against bacterial prolyl endopeptidase from Flavobacterium meningosepticum (IC50 25 microM). No effect of lipohexin (I) was found on the activity of mechanistically related proteases such as proline specific proteases and other serine proteases.

Animals↗
PubMed1997

[Pupillary diameter and pupillary reactions in heroin dependent patients and in patients participating in a methadone and morphine replacement program].

The computer-assisted static and dynamic light evoked pupillometry (TV-pupillometer 1050, Whittaker Corp.) had been proved to be a sensitive procedure for assessment of the effect of psychoactive drugs. Therefore, this method was used in 26 heroin dependent patients (mean age 24.42 years), 20 methadone substituted patients (mean age 29.75), and 20 morphine-substituted patients (mean age 30.65 years) to answer the question whether there were no differences within the patient groups but significant differences between the patients and healthy normals. Indeed, pupillary diameter (vegetative activation) as well as relative change (pupillary reagibility) showed no significant differences between the heroin dependents, the methadone substitution group and the morphine substitution group. However concerning pupillary diameter and relative change the patient groups differed significantly from the healthy controls. Onset latency revealed no differences within the patient groups and between patient groups and healthy controls respectively. Thus the variable pupillary diameter and relative change could be used to assess the additional application of opiates in patients participating in a substitution program.

Adult↗