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U Pfueller

Publications and source records attributed to U Pfueller.

12 recordsLinked to original sources
PubMed2005

Cloning and expression of mistletoe lectin III B-subunit.

Aqueous extracts of mistletoe (Viscum album L.) contain toxic proteins (lectins) MLI (viscumin), MLII, and MLIII. We previously cloned the gene encoding MLIII precursor. In the present study, a gene fragment encoding the carbohydrate-binding subunit of mistletoe toxic lectin MLIII was cloned and expressed in Escherichia coli cells. The structure and immunochemical properties of recombinant MLIII B-subunit were investigated using a panel of monoclonal antibodies against ML-toxins. Sugar-binding activity of recombinant MLIII B-subunit was determined by ELISA. Amino acid sequence analysis of the cloned MLIII compared with known mistletoe toxins and other ribosome-inactivating type II proteins (ricin, abrin a, and nigrin b B-subunits) revealed essential features of the recombinant MLIIIB primary structure that could determine sugar specificity of the lectin as well as immunomodulating and anti-tumor properties of mistletoe extracts.

Amino Acid Sequence↗
PubMed2004

Frontal and temporal sources of mismatch negativity in healthy controls, patients at onset of schizophrenia in adolescence and others at 15 years after onset.

Mismatch negativity (MMN) is an event-related potential measure of auditory change detection. It is widely reported to be smaller in patients with schizophrenia and may not improve along with otherwise successful clinical treatment. The main aim of this report is to explore ways of measuring and presenting four features of frequency-deviant MMN dipole sources (dipole moment, peak latency, brain location and orientation) and to relate these to the processes of psychopathology and illness progression. Data from early onset patients (EOS) at the start of the illness in adolescence, and others who had their first break in adolescence 15 years ago (S-15Y) were compared with two groups of age-matched healthy controls (C-EOS, C-15Y). A four-source model fitted the MMN waveform recorded from all four groups, whether MMN amplitude was more (EOS) or less (S-15Y) reduced. The locations were in the left superior temporal and anterior cingulate gyri, right superior temporal and inferior/mid frontal cortices. Dipole latencies confirmed a bottom-up sequence of processing and dipole moments were larger in the temporal lobes and on the left. Patients showed small dipole location changes that were more marked in the S-15Y than the EOS group (more rostral for the left anterior cingulate, more caudal for the right mid-frontal dipole) consistent with illness progression. The modelling of MMN dipole sources on brain atlas and anatomical images suggests that there is a degree of dissociation during illness between small progressive anatomical changes and some functional recovery indexed by scalp recordings from patients with an onset in adolescence 15 years before compared to adolescents in their first episode.

Adolescent↗
PubMed2004

Cloning and expression of catalytic subunit of MLIII, the ribosome-inactivating protein from Viscum album.

We have cloned the gene encoding a precursor of mistletoe (Viscum album) toxin MLIII. Analyses of nucleotide and deduced amino acid sequences of this gene revealed significant differences between MLI and MLIII preprotoxin genes. Immunochemical properties of recombinant A-subunit expressed in Escherichia coli and renatured were investigated using a panel of monoclonal antibodies raised against three mistletoe toxins (MLI, MLII, and MLIII). Ribosome-inactivating activity of recombinant MLIII A-subunit was detected in cell-free lysate of rabbit reticulocytes.

Amino Acid Sequence↗
PubMed1999

Study of heterogeneity of lectins in mistletoe preparations by monoclonal antibodies to their A-subunits.

Monoclonal antibodies (monAbs) displaying diverse affinity to native and recombinant mistletoe lectins A-chains (ML I-A, ML II-A, ML III-A and rML-A) have been obtained. In accordance to specificity of monAb they have been classified into three groups: 1. monAb against MLI-A and MLII-A, 2. monAb against MLII-A and MLIII-A, 3. monAb against A-subunits of MLI, MLII and MLIII. The results indicate, that antigen determinants of mistletoe lectins recognized by monAb MNA4, MNA9 and MTC12 do not contain any carbohydrates. Assay of lectins in mistletoe preparations was based on enzyme-linked lectin assay to meet the requirements given in the guidelines for drug tests. In this paper a sandwich ELISA test-system is described which allows to identify each of three ML-toxins. With the detection limits below 3 ng/ml and a linear measuring range of 3-30 ng/ml, dosages of mistletoe lectins in therapeutic range can be assayed. The problems in mistletoe lectins determination, structural differences and nature of heterogeneity of this proteins are discussed.

Animals↗
PubMed1998

Mistletoe lectin I forms a double trefoil structure.

The quaternary structure of mistletoe lectin I (MLI), a type II ribosome inactivating protein, has been determined by X-ray crystallography. A definitive molecular replacement solution was determined for MLI using the co-ordinates of the homologue ricin as a search model. MLI exists as an [AB]2 dimer with internal crystallographic two-fold symmetry. Domain I of the B chains is non-covalently associated through interactions involving three looped chains (alpha, beta, gamma) in each molecule of the dimer, forming a double trefoil structure. The ricin molecule which shares 52% sequence homology with MLI has a disulphide bridge between Cys20 and Cys39 in the alpha loop. An evolutionary mutation has replaced Cys39 with serine in MLI. This mutation appears to allow the alpha loop the flexibility required to take up its place at the dimer interface, and also suggests a rationale for why ricin does not form dimers. Measurement of retention times using FPLC gel filtration confirms that dimerisation also occurs in solution between MLI B chains with an association constant Ka = 10(6) M.

Amino Acid Sequence↗
PubMed1997

Testing the hypothesis of the relationships between laterality and ability according to Annett's right-shift theory: findings in an epidemiological sample of young adults.

In a large epidemiological sample of young adults, predictions of the right-shift (RS) theory of Annett that cognitive abilities will vary with right-left hand skill were tested. Presenting a theory of a genetic balanced polymorphism with heterozygote advantage for laterality and ability, Annett & Manning (1989, 1990a) and Annett (1993c) claimed that probands at the right end of the R-L hand skill continuum would show lower general intelligence in IQ testing and that specific verbal abilities and educational success would be lower at both extremes of the R-L distribution, taking the form of an inverted U. Most of these predictions could not be confirmed by our study. In particular, our data contrast with the important and specific prediction of the RS theory that strong dextrals will be the most disfavoured group. In our sample, probands at the left end of the R-L continuum had significantly lower scores in spelling and educational success and showed a tendency to have lower non-verbal IQ scores, while strong dextrals tended to have average or even marginally higher ability scores. The effects, however, are small and decrease when controlling for other variables. Implications of these empirical findings for the right-shift theory are discussed.

Achievement↗
PubMed1995

The interactions of anti-MLI monoclonal antibodies with isoforms of the lectin from Viscum album.

Monoclonal antibodies (mAb) reacting with native (TA5, TB12) and denatured (T33, T35) plant toxin mistletoe lectin I (MLI) from Viscum album have been obtained. The interaction between mAbs and native toxin with ML isoforms (MLII, MLIII) has been investigated. An immunological cross-reaction has been shown to take place for mAb TA5 (anti-A-chain of MLI) between MLII and MLIII isoforms of toxin. TA5 has not inhibited enzyme activity of the A-chain in a rabbit reticulocyte cell-free system. TB12 has been shown to react with the galactose-binding site of the B-chain. TA5 and TB12 have shown no cross-reaction with plant toxin ricin. The association constants for mAbs have been determined. The nature of heterogeneity of the lectins from Viscum album is discussed.

Animals↗
PubMed1994

Renaturated ricin toxin B chain made in Escherichia coli is soluble, stable, and biologically active.

Escherichia coli cells transformed with plasmids containing ricin B-chain coding sequences are shown to express this heterologous protein in inclusion bodies. After denaturation and renaturation of the product in the presence of glutathione and lactose, the recombinant ricin B-chain is soluble, biologically active and stable. Cytotoxicity of heterodimer with this protein and ricin A-chain is bound to be only ten times less than of native ricin. Recombinant B-chain alone was nontoxic to cells (ID50 > 10(-6)M). Our data suggest that N-glycosylation of ricin B-chain is not required for its biological activity.

Cell Line↗
PubMed1993

Comparison of the cytotoxic activity of the immunotoxins with different internalization rate.

In comparative experiments the conjugates of A-chain of a plant toxin ricin (RTA) and monoclonal antibody (MAb) HAE9 (IgM) directed against human erythroblast antigen (Ag-Eb) or Mab HAE3 against human glycophorin-A and immunotoxins (IT) directed to CD5 and CD7 antigens of human T-lymphocytes have been investigated. Proceeding from the number of receptors on a cell, we compared efficiency of the cytotoxic effects of the conjugates with different internalization rate. The efficiency of immunoconjugates against Ags with an extremely high internalization rate was only slightly higher than that of immunoconjugates against Ags with a lower internalization rate. The enhancing effect of ammonium chloride on immunoconjugate cytotoxicity was more pronounced in the case of immunotoxins with a high internalization rate.

Antibodies, Monoclonal↗
PubMedDate not supplied

[Production of biologically active recombinant ricin B-chain].

Escherichia coli cells transformed with plasmids containing ricin B-chain coding sequences are shown to express this heterologous protein in inclusion bodies. After denaturation and renaturation of the product in the presence of glutathione and lactose, the recombinant ricin B-chain is soluble, biologically active and stable. Cytotoxicity of heterodimer containing this protein and ricin A-chain is found to be only ten times lower, than that of native ricin. Recombinant B-chain alone was nontoxic to cells (ID50 > 10(-6) M). Our data suggest that ricin B-chain oligosaccharides are essential for stability preserving protein from proteolytic degradation in cells.

Base Sequence↗