Organ toxicity and quality of life after allogeneic bone marrow transplantation in pediatric patients: a single centre retrospective analysis.

One hundred and fifty five pediatric patients underwent allogeneic bone marrow transplantation between 1980 and 1996 in the St Anna Children's Hospital in Vienna with an overall survival of 52.3% (81 patients). Seventy-three patients with a minimum observation time of 1 year (1-13 years, median: 4.

Engineering activity and stability of Thermotoga maritima glutamate dehydrogenase. II: construction of a 16-residue ion-pair network at the subunit interface.

The role of an 18-residue ion-pair network, that is present in the glutamate dehydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus, in conferring stability to other, less stable homologous enzymes, has been studied by introducing four new charged amino acid residues into the subunit interface of glutamate dehydrogenase from the hyperthermophilic bacterium Thermotoga maritima. These two GDHs are 55 % identical in amino acid sequence, differ greatly in thermo-activity and stability and derive from microbes with different phylogenetic positions. Amino acid substitutions were introduced as single mutations as well as in several combinations.

The catalytic reaction and inhibition mechanism of Drosophila alcohol dehydrogenase: observation of an enzyme-bound NAD-ketone adduct at 1.4 A resolution by X-ray crystallography.

Drosophila alcohol dehydrogenase (DADH) is an NAD+-dependent enzyme that catalyzes the oxidation of alcohols to aldehydes/ketones. DADH is the member of the short-chain dehydrogenases/reductases family (SDR) for which the largest amount of biochemical data has been gathered during the last three decades. The crystal structures of one binary form (NAD+) and three ternary complexes with NAD+.

Refined crystal structure of a superoxide dismutase from the hyperthermophilic archaeon Sulfolobus acidocaldarius at 2.2 A resolution.

The extremely thermostable superoxide dismutase from the hyperthermophilic archaeon Sulfolobus acidocaldarius was crystallized and the three-dimensional structure was determined by X-ray diffraction methods. The enzyme crystallized in the monoclinic spacegroup C2 with the cell dimensions a=168.1 A, b=91.

Proteins from thermophilic and mesophilic organisms essentially do not differ in packing.

The role of the packing density in the elevation of thermal stability of proteins from thermophilic organisms is widely discussed in the literature. In the present study, this issue was reconsidered in the scale of an unbiased set of protein structures. Partial specific volumes, void and cavity volumes were calculated for a set of 80 non-homologous proteins and for 24 proteins from thermophilic organisms and analysed in the context of their possible role in thermal stabilization.

Two-dimensional crystallization of the chaperonin TF55 from the hyperthermophilic archaeon Sulfolobus solfataricus.

Oligomers of the chaperonin TF55 from Sulfolobus solfataricus have been successfully crystallized in two dimensions via their interaction with a phospholipid monolayer at the air/liquid interface. Oligomer orientation was dependent upon the lipid headgroup used. A neutral lipid monolayer gave rise to small paracrystalline areas of TF55 side views, whereas a negatively charged lipid monolayer resulted in large coherent crystalline areas of the chaperonin in an end-on orientation.

The refined crystal structure of Drosophila lebanonensis alcohol dehydrogenase at 1.9 A resolution.

Drosophila alcohol dehydrogenase (DADH; EC 1.1.1.

Transplantation activities and treatment strategies in paediatric stem cell transplantation centres: a report from the EBMT Working Party on Paediatric Diseases. European Group for Blood and Marrow Transplantation.

To determine the current approach to stem cell transplantation (SCT) in centres which treat predominantly paediatric patients, a questionnaire was sent to 67 centres known by the EBMT registry to perform SCT mainly in children. Fifty-five centres from 19 countries responded. Forty centres (75%) started their transplantation activities between 1980 and 1992.

Isolation and structure of repressor-like proteins from the archaeon Sulfolobus solfataricus. Co-purification of RNase A with Sso7c.

The thermostable histone-like protein Sso7c (Sso for Sulfolobus solfataricus) from the archaeon Sulfolobus solfataricus was purified from the supernatant of acid-soluble cell lysates. Reverse phase HPLC of an apparently homogeneous Sso7c protein fraction from Mono S chromatography resulted in resolution of three further peaks. Sequence analysis revealed one of these components to be bovine RNase A, originating from the culture medium and explaining the RNA hydrolyzing activities of Sso7 preparations previously described.

Resolution of early cytomegalovirus (CMV) infection after leukocyte transfusion therapy from a CMV seropositive donor.

A 2 year and 8 month old CMV-negative boy suffering from stage III neuroblastoma underwent ABMT in first very good partial remission. He acquired early CMV infection on day +5, followed by consecutive graft failure and severe sepsis, and the clinical course deteriorated. Between days +16 and +21, he received seven leukocyte concentrates (LC) collected from a healthy, but CMV-IgG-seropositive relative stimulated with G-CSF (filgastrim, 5 microg/kg/day).

Proteins from hyperthermophiles: stability and enzymatic catalysis close to the boiling point of water.

It has become clear since about a decade ago, that the biosphere contains a variety of microorganisms that can live and grow in extreme environments. Hyperthermophilic microorganisms, present among Archaea and Bacteria, proliferate at temperatures of around 80-100 degrees C. The majority of the genera known to date are of marine origin, however, some of them have been found in continental hot springs and solfataric fields.

A protein disulfide oxidoreductase from the archaeon Pyrococcus furiosus contains two thioredoxin fold units.

Protein disulfide bond formation is a rate limiting step in protein folding and is catalyzed by enzymes belonging to the protein disulfide oxidoreductase superfamily, including protein disulfide isomerase (PDI) in eucarya and DsbA in bacteria. The first high resolution X-ray crystal structure of a protein disulfide oxidoreductase from the hyperthermophilic archaeon Pyrococcus furiosus reveals structural details that suggest a relation to eukaryotic PDI. The protein consists of two homologous structural units with low sequence identity.

Architecture of nonspecific protein-DNA interactions in the Sso7d-DNA complex.

Many biochemical processes, including DNA packing, maintenance and control, rely on non-sequence specific protein-DNA interactions. Nonspecific DNA-binding proteins have evolved to tolerate a wide range of DNA sequences, yet bind with a respectable affinity. The nonspecific binding requirement is in contrast to that imposed on, for example, transcription factors and implies a different structural basis for the biomolecular recognition process.

Engineering activity and stability of Thermotoga maritima glutamate dehydrogenase. I. Introduction of a six-residue ion-pair network in the hinge region.

Comparison of the recently determined three-dimensional structures of several glutamate dehydrogenases allowed for the identification of a five-residue ion-pair network in the hinge region of Pyrococcus furiosus glutamate dehydrogenase (melting temperature 113 degrees C), that is not present in the homologous glutamate dehydrogenase from Thermotoga maritima (melting temperature 93 degrees C). In order to study the role of this ion-pair network, we introduced it into the T. maritima enzyme using a site-directed mutagenesis approach.

CMV-viraemia during allogenic bone marrow transplantation in paediatric patients: association with survival and graft-versus-host disease.

Sixty-one consecutive paediatric patients undergoing allogenic bone marrow transplantation (BMT) were screened prospectively for cytomegalovirus (CMV)-viraemia by PCR. Sixteen patients (26%) presented with single or recurrent CMV-viraemia between day -7 and + 100. Although only four of them had evidence of CMV-disease, there was a significant difference in the incidence of acute Graft versus Host Disease (GVHD) grade III-IV (75% vs 15.

In vitro DNA binding of the archaeal protein Sso7d induces negative supercoiling at temperatures typical for thermophilic growth.

The topological state of DNA in hyperthermophilic archaea appears to correspond to a linking excess in comparison with DNA in mesophilic organisms. Since DNA binding proteins often contribute to the control of DNA topology by affecting DNA geometry in the presence of DNA topoisomerases, we tested whether the histone-like protein Sso7d from the hyperthermophilic archaeon Sulfolobus solfataricus alters DNA conformation. In ligase-mediated supercoiling assays carried out at 37, 60, 70, 80 and 90 degrees C we found that DNA binding of increasing amounts of Sso7d led to a progressive decrease in plasmid linking number (Lk), producing negative supercoiling.

Thermal unfolding of small proteins with SH3 domain folding pattern.

The thermal unfolding of three SH3 domains of the Tec family of tyrosine kinases was studied by differential scanning calorimetry and CD spectroscopy. The unfolding transition of the three protein domains in the acidic pH region can be described as a reversible two-state process. For all three SH3 domains maximum stability was observed in the pH region 4.

Evidence for local 32 symmetry in homotrimeric riboflavin synthase of Escherichia coli.

Riboflavin synthase is a trimer of identical 23-kDa subunits. The primary structure is characterized by considerable similarity of the C-terminal and N-terminal parts. Recombinant riboflavin synthase of Escherichia coli and Bacillus subtilis was crystallized by the vapor diffusion method.

Thermodynamic characterization of non-sequence-specific DNA-binding by the Sso7d protein from Sulfolobus solfataricus.

We used isothermal titration calorimetry and fluorescence spectroscopy to investigate the thermodynamics of non-sequence-specific DNA-binding by the Sso7d protein from the archaeon Sulfolobus solfataricus. We report the Sso7d-poly(dGdC) binding thermodynamics as a function of buffer composition (Tris-HCl or phosphate), temperature (15 to 45 degrees C), pH (7.1 to 8.

1070 myeloablative megatherapy procedures followed by stem cell rescue for neuroblastoma: 17 years of European experience and conclusions. European Group for Blood and Marrow Transplant Registry Solid Tumour Working Party.

1070 myeloablative procedures followed by stem cell rescue for neuroblastoma are reviewed. These 1070 procedures are part of the European Group for Blood and Marrow Transplant (EBMTG) registry from the last 17 years (in 4536 patients). In 1070 neuroblastoma patients, survival at 2 years was 49%, at 5 years, 33% and relapses were observed as late as 7 years post-BMT (bone marrow transplant).

Multivariate analysis of risk factors in stage 4 neuroblastoma patients over the age of one year treated with megatherapy and stem-cell transplantation: a report from the European Bone Marrow Transplantation Solid Tumor Registry.

Purpose: The European Bone Marrow Transplantation (EBMT) Solid Tumor Registry (STR) contains detailed information on children with advanced neuroblastoma who, after standard-dose induction chemotherapy and surgery, received myeloablative megatherapy (MGT) followed by stem-cell transplantation (SCT). This data base was analyzed to identify factors that predict event-free survival (EFS).

Patients And Methods: Eligibility criteria were stage IV neuroblastoma, age over 1 year at diagnosis, and no relapse before MGT/SCT.

Predictive potential of testing for bone marrow involvement in Ewing tumor patients by RT-PCR: a preliminary evaluation.

EWS/ets-oncogene fusion transcripts can be detected in at least 98% of Ewing tumors [(ET) Ewing sarcoma and peripheral primitive neuroectodermal tumor] by reverse transcriptase-polymerase chain reaction (RT-PCR), thus confirming the histopathologic diagnosis. To detect minimal amounts of tumor cells in the bone marrow (BM), we used an RT-PCR assay with a high sensitivity, revealing one tumor cell in a background of 10(6) normal cells. We examined BM samples from 35 newly diagnosed ET patients (23 with localized and 12 with metastatic disease).

The present role of bone marrow and stem cell transplantation in the therapy of children with acute leukemia.

Hematopoetic stem cell transplantation (SCT) often represents a unique opportunity for curing children with leukemia. Nevertheless, selecting the patient who could really benefit from this procedure remains a controversial issue. The current consensus is as follows: About 20% of children with ALL can be defined as high-risk patients by criteria such as t(9;22), t(4;11), no complete remission at day 42, poor prednisone response, and T-immunophenotype or pre-pre B-ALL, myeloid markers or more than 100,000 white blood cells/microliter.

High-dose chemotherapy with autologous bone marrow rescue in children with poor-risk Burkitt's lymphoma: a report from the European Lymphoma Bone Marrow Transplantation Registry.

To evaluate the role of high-dose chemotherapy (HDC) followed by autologous bone marrow transplantation (ABMT) in children with poor-prognosis Burkitt's lymphoma, the European Lymphoma BMT registry was critically reviewed. Between February 1979 and July 1991, a selected group of 89 children (78 boys and 11 girls) were considered as ABMT candidates in 12 European cancer centers for the following reasons: poor initial response (PIR) to first-line chemotherapy in 28 patients, primary refractory disease (PRD) in nine patients, sensitive relapse (SR) in 38 patients, and resistant relapse (RR) in 14 patients. The median age at ABMT was 8.

Drosophila alcohol dehydrogenase: evaluation of Ser139 site-directed mutants.

Drosophila alcohol dehydrogenase (DADH) belongs to the large and highly heterogeneous (15-30% residue identity) short-chain dehydrogenase/reductase family (SDR). It is the only reported member that oxidizes mainly ethanol and 2-propanol among other alcohols. To confirm the role of Ser139 we constructed two site-directed mutants, Ser139Ala and Ser139Cys, which show no enzymatic activity.