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Biomedical subjects

W Machleidt

Publications and source records attributed to W Machleidt.

At least 55 records · Page 3Linked to original sources
PubMed1991

[The dynamics of emotional states in patients with epilepsy. A phenomenologic EEG analysis study].

The history of the main ideas of psychogenetic epilepsy-theory is outlined. Using the antithesis of omnipotency and impotency further implications of this theoretical approach are explained. Epileptic patients suffer from a deficient structure of their self and do not overcome infantile obsolutistic thought, behavioral and emotional patterns. Their behavior and symptoms exhibit the characteristics of the "all or nothing schema". In particular conflict solving strategies of epileptics consist either in an exaggerated spirit of toleration or in violent outbursts of aggression. Destructive fits of rage, epileptic fits and psychotic derangements can be understood as manifestations of aggression on ontogenetically earlier developmental stages. From this point of view we studied the influence of the emotional states of epileptics on EEG-background activity and discharges: Conflict-centered interviews were recorded simultaneously in ten epileptic patients with EEG- and video-techniques. FFT-spectralanalysed EEG-data and discharges from visually evaluated EEG-records were correlated with hermeneutic analyses and the ratings of the emotional states. Results show significant correlations between spectral EEG-parameters and aggression, anxiety, sorrow and joy. These results were validated by further studies. Normal aggressive episodes were correlated with an increase of power in all frequency-bands. Epileptic discharges occurred in over or latent aggressive impulses. In the theory of the "Kontinuitätsparadigma" a parallelism of two transitional stages is suggested: ranging from normal to pathological aggression with all its symptoms and from EEG-dynamics in normal aggression to typical discharges in epileptic aggression. The latter represents the extreme pathological end of the stage while the former is its starting-point.

Adult↗
PubMed1991

A triazolodiazepine platelet activating factor receptor antagonist (WEB 2086) reduces pulmonary dysfunction during endotoxin shock in swine.

We wanted to determine the effects of WEB 2086, a platelet activating factor (PAF) antagonist, in lipopolysaccharide (LPS) shock in anesthetized pigs. In a randomized study, LPS from S. abortus equi, 2 micrograms/kg/h was given IV for six hours. Thirteen animals received LPS and WEB 2086, 10 mg/kg/h IV for 6.5 hours, beginning 30 minutes before LPS. Eleven septic controls received saline and LPS, three nonseptic controls received saline and WEB 2086, and three nonseptic controls received saline only. In six animals we investigated the effect of synthetic PAF in doses between 50 and 10,000 ng on arterial (AP) and pulmonary arterial (PAP) pressure before and during infusion of WEB 2086. The LPS-induced rise in PAP was reduced by WEB 2086 (p = 0.01) but not the decrease in AP. The LPS-induced leukopenia, hypoxia, increase in airway pressure, and release of plasminogen activator inhibitor were reduced by WEB 2086. Platelet activating factor produced an increase in PAP and a biphasic response in AP. All PAF dose response curves were shifted to the right by WEB 2086. Platelet activating factor was a pulmonary hypertensive agent and contributed to the LPS-induced respiratory alterations.

Animals↗
PubMed1991

Possible role of extracellularly released phagocyte proteinases in coagulation disorder during liver transplantation.

Orthotopic liver transplantation is frequently associated with a complex coagulation disorder, influencing the outcome of the procedure. In this respect, disseminated intravascular coagulation (DIC) had been suggested to be of causative importance for bleeding complications after reperfusion of the liver graft. In 10 consecutive patients undergoing orthotopic liver transplantations, we studied the occurrence of two phagocyte proteinases of different origin in the graft liver perfusate and in systemic blood during the operation, as well as their effects on hemostasis. As compared with plasma samples taken at the end of the anhepatic phase, highly significant increases of cathepsin B and thrombin-anti-thrombin III complexes (TAT), as well as highly significant decreases in antithrombin III, protein C, and C1-inhibitor were observed in graft liver perfusate. Von Willebrand factor and fibrinogen were slightly decreased, whereas the elastase-alpha 1 proteinase inhibitor complexes (EPI) were elevated. In plasma the activity of cathepsin B remained unchanged during the prereperfusion phases, but immediately after revascularization of the graft this cysteine proteinase increased. The EPI showed a gradual increase in plasma during the preanhepatic and anhepatic phases but a more pronounced increase in the reperfusion phase. In parallel with the rise in these two proteinases TAT increased and the activities of antithrombin III and C1-inhibitor in plasma decreased after reperfusion. At 12 hr after revascularization plasma levels of TAT, antithrombin III, and C1-inhibitor had returned to the prereperfusion ranges, whereas cathepsin B and EPI were significantly above the baseline levels. These observations are consistent with the hypothesis that extracellularly released lysosomal proteinases may play a role in the development of a DIC-like constellation, including thrombin formation after revascularization of the liver graft. For the first time we could prove the occurrence of phagocyte proteinases in graft liver perfusate and evaluate the importance of these proteinases for the understanding of the pathophysiology leading to bleeding complications in patients undergoing orthotopic liver transplantation.

Antithrombin III↗
PubMed1991

Proteolysis of defensive proteins in peritonitis exudate: pathobiochemical aspects and therapeutical approach.

Peritonitis exudate reveals strong proteolytic activity which is paralleled by deficient opsonic capacity and high concentrations of lysosomal proteinases (elastase and cathepsin B). Lysosomal serine and cysteine proteinases (elastase, cathepsins B, L) were shown to degrade immunoglobulin G(IgG) and seem to be at least partially responsible for the observed proteolytic inactivation of IgG in peritonitis exudates. Intraabdominal serum application seems to restore opsonic activity by substitution of opsonins and proteinase inhibitors.

Ascitic Fluid↗
PubMed1991

Molecular mechanism of inhibition of cysteine proteinases by their protein inhibitors: kinetic studies with natural and recombinant variants of cystatins and stefins.

Natural and recombinant variants of the cysteine proteinase inhibitors chicken cystatin and human stefin B were characterized by determination of their inhibition constants for papain, actinidin and human cathepsins B and H. The individual contributions of the three contact regions to the binding energy of the chicken cystatin-papain complex were calculated as 36% for the N-terminal segment, 51% for the first and 13% for the second hairpin loop. Removal of the N-terminal contact region of chicken cystatin resulted in a 10000-fold lower affinity for papain. In contrast, stefin B remained a tight-binding inhibitor of papain and actinidin without its N-terminal segment. Affinity of stefin B for papain was only slightly affected by exchange of the residue predicted to bind in the S2 subsite of papain. The essential contribution of the first hairpin loop to inhibitor binding was confirmed by the 240-fold lower affinity for papain of a Val48----Asp mutant of stefin B. Inhibition of cathepsin B by stefins A and B is slow-binding. Binding of stefin B, not of stefin A, follows a two-step mechanism involving a slow isomerisation of the enzyme-inhibitor complex.

Amino Acid Sequence↗
PubMed1990

The complete amino-acid sequences of human ganglioside GM2 activator protein and cerebroside sulfate activator protein.

The complete amino-acid sequences of human ganglioside GM2 activator protein and cerebroside sulfate activator protein have been established by Edman degradation. The GM2 activator is composed of 162 amino acids, the first two serine residues being present in only 20% of the material. A single carbohydrate chain is N-glycosidically linked to Asn32. Three hydrophobic alpha-helices may contribute to its lipid-binding site. Three amino acids differ from those found by cDNA sequencing which may be due to a polymorphism. The cerebroside sulfate activator consists of 80 amino acids and carries one N-linked carbohydrate chain at Asn21. The C-terminal valine residue is lacking in about 80% of the material. In spite their similar functions, both activator proteins show no sequence or structural similarities.

Amino Acid Sequence↗
PubMed1990

[Coordination and planning in community psychiatry. 14-year-experience in Hannover between the recommendations of the Psychiatric Inquiry and the Expert Commission (1976-1989)].

Against the background of the model pattern conceived in West Germany in 1975 on the basis of the so-called "Psychiatrie-Enquête" and in 1988 on recommendations made by a committee of experts, the article describes and discusses the history, composition, manner of working, thematic scope and the results of the coordination round table concerning psychiatric care under community council auspices. This model has been realised in Hanover since 1976 as part of the segmentation of psychiatric care according to community districts. The study aims at stimulating suggestions for a general solution regarding coordination- and planning of communal psychiatric care and how to translate these suggestions into reality, both on a scientific basis and with due reference to the realities of the prevailing circumstances. This can help to avoid developments in a wrong direction and also to avoid delays that may arise in solving this core problem of sociopsychiatric reform.

Community Mental Health Services↗
PubMed1989

Mechanism of inhibition of papain by chicken egg white cystatin. Inhibition constants of N-terminally truncated forms and cyanogen bromide fragments of the inhibitor.

N-terminally truncated forms of chicken egg white cystatin and its cyanogen bromide fragments were isolated and assayed for inhibition of papain. Truncated forms beginning with Gly-9 and Ala-10 had a 5000-fold lower affinity for papain than the two isoelectric forms (pI = 6.5 and 5.6) of the full-length inhibitor (Ki = 6 pM and 7 pM) or a truncated form beginning with Leu-7 (Ki = 6 pM), indicating the outstanding importance of one or two residues preceding conserved Gly-9 for binding. A weak inhibition of papain (Ki = 900 nM) was exhibited by the intermediate cyanogen bromide fragment (residues 30-89) containing the chicken cystatin QLVSG variation of the QVVAG segment which is conserved in almost all members of the cystatin superfamily. The obtained affinity data provide independent evidence for the validity of the proposed docking model of a chicken cystatin-papain complex [(1988) EMBO J. 7, 2593-2599].

Amino Acid Sequence↗
PubMed1988

Proteinase-sensitive regions in the heavy chain of low molecular weight kininogen map to the inter-domain junctions.

Low molecular weight kininogen from human plasma was subjected to limited proteolysis with trypsin, chymotrypsin, elastase, and bromelain, and the resulting fragments of 20,000 or 40,000 Da were isolated. Amino-terminal sequence analysis of the fragments disclosed for the various proteinases eight independent cleavage sites distinct from the typical kallikrein cleavage sites flanking the kinin region. All the identified cleavage sites cluster in two stretches of 11-12 residues of the kininogen heavy chain. These short segments represent the primary attack sites for proteinases ("proteinase-sensitive regions") in the heavy chain portion of human low molecular weight kininogen. The amino acid sequences of the two proteinase-sensitive regions are mutually homologous; they are further characterized by the presence of a single copy each of the consensus tetrapeptide Cys-X-Gly-Cys known to form a narrow disulfide loop (Kellermann, J., Thelen, C., Lottspeich, F., Henschen, A., Vogel, R., and Müller-Esterl, W. (1987) Biochem. J. 247, 15-21). The proteinase-sensitive regions are located at the junctions of the three cystatin-like domains constituting the kininogen heavy chain. Proteolytic cleavage at the sensitive regions dissects the kininogen heavy chain and releases single domains of 20,000 Da and combined domains of 40,000 Da which can function as cysteine proteinase inhibitors. The presence of kininogen heavy chain domains in plasma samples under pathologic conditions suggests that cleavage of the proteinase-sensitive regions might also occur in vivo.

Amino Acid Sequence↗
PubMed1988

Amino acid sequences of the human kidney cathepsins H and L.

The complete amino acid sequences of human kidney cathepsin H (EC 3.4.22.16) and human kidney cathepsin L (EC 3.4.22.15) were determined. Cathepsin H contains 230 residues and has an Mr of 25116. The sequence was obtained by sequencing the light, heavy and mini chain and the peptides produced by cyanogen bromide cleavage of the single-chain form of the enzyme. The glycosylated mini chain is a proteolytic fragment of the propeptide of cathepsin H. Human cathepsin L has 217 amino acid residues and an Mr of 23720. Its amino acid sequence was deduced from N-terminal sequences of the heavy and light chains and from the sequences of cyanogen bromide fragments of the heavy chain. The fragments were aligned by comparison with known sequences of cathepsins H and L from other species. Cathepsins H and L exhibit a high degree of sequence homology to cathepsin B (EC 3.4.22.1) and other cysteine proteinases of the papain superfamily.

Amino Acid Sequence↗
PubMed1988

The precursor of sulfatide activator protein is processed to three different proteins.

The enzymic degradation of a number of sphingolipids in the lysosomes is stimulated by small acid glycoproteins named activator proteins. We purified and sequenced a new protein, called component C, which seems to be related to sulfatide activator and to a recently described activator of glucosylceramidase (A1 activator) (Kleinschmidt, T., Christomanou, H. & Braunitzer, G. (1987) Biol. Chem. Hoppe-Seyler 368, 1571-1578). It consists of 78 amino acids and carries one carbohydrate chain at aparagine 20. Component C shows 21.5% sequence homology to sulfatide activator and 34.2% homology to A1 activator. Structural similarities between these three proteins have also been detected. Recently the cDNA sequence of the sulfatide activator precursor has been published (Dewji, N.N., Wenger, D.A. & O'Brien, J.S. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 8652-8656). We could align the protein sequences of sulfatide activator, A1 activator and component C with that of this large precursor protein. After minor corrections of the DNA sequence we obtained total fit. Thus it seems that three different proteins are derived from the sulfatide activator precursor by proteolytic processing. Possible processing sites were found on the precursor at sites adjacent to the N-termini and C-termini of the mature proteins. The processing of sulfatide activator was studied by Fujibayashi and Wenger (Fujibayashi, S. & Wenger, D.A. (1986) Biochim. Biophys. Acta 875, 554-562). Their data support our assumption that processing occurs by simultaneous cleavage at all possible sites.

Amino Acid Sequence↗
PubMed1988

Molecular cloning and sequencing of a cDNA coding for mature human kidney cathepsin H.

A cDNA library was established from human kidney RNA and screened with an extended oligonucleotide probe derived from the amino-acid sequence of human cathepsin H. A recombinant clone, pRF15, was isolated and characterized. DNA sequence analysis of its 1106-nucleotide-long insert revealed that pRF15 encodes the complete protein sequence of mature cathepsin H plus 28 amino acids of a propeptide, thus confirming that cathepsin H is synthesized as a larger precursor molecule and posttranslationally processed. Northern blot analysis indicated that cathepsin H is predominantly synthesized in kidney. A high degree of sequence homology was observed with rat cathepsin H, especially within the propeptide. The part of the prosequence coding for the "minichain" is conserved in the prosequence of aleurain, a plant thiol protease.

Amino Acid Sequence↗
PubMed1988

Inhibitorily active recombinant human stefin B. Gene synthesis, expression and isolation of an inhibitory active MS-2 pol-stefin B fusion protein and preparation of Des[Met1,2(2)]stefin B.

A synthetic gene coding for the human intracellular cysteine proteinase inhibitor, stefin B, was constructed from 13 chemically synthesized oligonucleotides according to the method of Khorana. The gene was inserted into the plasmid vector pTZ, amplified and sequenced. For expression, a temperature-inducible system producing fusion proteins was used. With the vector pEx31A containing the synthetic cystatin B gene, E. coli strain 537 produced a fusion protein of the N-terminal part of bacteriophage MS-2 polymerase and [Met-2Gly-1]stefin B. Lysates of the induced bacteria were inhibitorily active against papain. The fusion protein was expressed in high yield (about 20% of total E. coli proteins) and mostly deposited as inclusion bodies. The unfolded fusion protein was partially purified in the presence of urea. After refolding, approx. 6% of the protein was inhibitorily active against papain, human cathepsin H and B. Des[Met1,2(2)]stefin B was released by cyanogen bromide cleavage of the fusion protein and identified by N-terminal amino-acid sequence analysis. The non-separated cleavage products were also inhibitorily active after refolding. The estimated inhibition constants for the fusion protein and its cleavage products were similar to those reported for natural stefin B.

Amino Acid Sequence↗
PubMed1988

Chemical synthesis of a gene for human cystatin C and its expression in E. coli.

A DNA containing the coding sequence for the human cysteine proteinase inhibitor protein cystatin C has been obtained by enzymatic ligation of chemically synthesized deoxyoligonucleotides, using the Khorana ligation method. The 375 bp synthetic gene carries signals for the translation initiation and termination and was expressed in E. coli as a beta-galactosidase fusion protein as well as a secreted protein under the control of the E. coli alkaline phosphatase signal sequence. The secreted hybrid protein was shown to have similar biological properties as the authentic protein isolated from human plasma.

Amino Acid Sequence↗
PubMed1988

A new purification procedure of human kidney cathepsin H, its properties and kinetic data.

A purification procedure of cathepsin H from human kidney is presented. It includes gel filtration, ion exchange chromatography, and covalent chromatography on thiol Sepharose as an essential step. Purified cathepsin H emerges in an isoelectric focusing gel at pH 6.1 and 6.3. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate shows a molecular mass of about 28 kDa. Less than 20% of the enzyme preparation can be separated into a heavy (24 kDa) and a light chain (4 kDa) after reduction and gel filtration on Sephacryl S-200. The partial amino-acid sequence of human cathepsin H shows its close similarity to rat cathepsin H. Inhibition constants (Ki) of cathepsins H and B with chicken cystatin, two forms of human stefin A, human stefin B, and two forms of human cystatin C are in the range of 10(-9) to 10(-11)M.

Amino Acid Sequence↗