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G Grabowski

Publications and source records attributed to G Grabowski.

14 recordsLinked to original sources
PubMed2003

Gaucher's disease: identification of novel mutant alleles and genotype-phenotype relationships.

A sequencing protocol for the acid beta-glucosidase (GCase) gene (GBA) was developed using a long-range PCR template. This protocol has an advantage of greater DNA yields over similar strategies. Seven Gaucher's disease patients had four novel and five other rare alleles. A non-pseudogene in-frame deletion (g.2600-2602delTAC) and a new complex mutation (null allele) were identified in Gaucher's disease type 1, i.e. the g.2600-2602delTAC deletion is associated with the non-neuronopathic variant. An F251L allele was found in a baby with the collodion skin phenotype. Three mutant alleles were identified in a single primary family with type 3. The patients' father at 45 years is healthy and is heteroallelic for the G202R and E326K alleles. Family studies indicated that E326K is in trans to G202R and L444P, and that isolated E326K is non-pathogenic in this family. A rare mutation R257Q was identified in a type 2 patient, providing an association with neuronopathic disease. A genotype L444P/L444P was noted in a 22-year-old non-neuronopathic patient. Complete gene sequencing showed a new complex allele consisting of L444P and g.7741T > C in the 3' UTR. Three additional complex alleles also involved the 3' UTR. Complete gene characterization in Gaucher's disease should allow greater insights into the correlation of specific alleles with phenotype.

Adolescent↗
PubMed1999

Combined genetic and physical map of the complex genome of Agrobacterium tumefaciens.

A combined genetic and physical map of the Agrobacterium tumefaciens A348 (derivative of C58) genome was constructed to address the discrepancy between initial single-chromosome genetic maps and more recent physical mapping data supporting the presence of two nonhomologous chromosomes. The combined map confirms the two-chromosome genomic structure and the correspondence of the initial genetic maps to the circular chromosome. The linear chromosome is almost devoid of auxotrophic markers, which probably explains why it was missed by genetic mapping studies.

Agrobacterium tumefaciens↗
PubMed1997

Enzyme therapy in Gaucher disease type 1: effect of neutralizing antibodies to acid beta-glucosidase.

Gaucher disease type 1, a non-neuronopathic lysosomal storage disease, is caused by mutations at the acid beta-glucosidase locus. Periodic infusions of macrophage-targeted acid beta-glucosidase reverse hepatosplenomegaly, hematologic, and bony findings in many patients. Two patients receiving enzyme therapy developed neutralizing antibodies to acid beta-glucosidase that were associated with a lack of improvement or progressive disease. After initial improvement, case 1 had no additional response to 2 years of high-dose (50 U/kg every 2 weeks) enzyme therapy. Similarly, case 2 initially showed a favorable response to enzyme therapy that plateaued after 1 year of treatment. Both patients developed minor allergic reactions and antibodies to acid beta-glucosidase within the first 6 months of treatment. Enzyme therapy was discontinued in case 1, with resultant disease progression and need for splenectomy. An immunosuppression/tolerization protocol was initiated in case 2 because of disease progression and stable neutralizing antibody titers. The IgG neutralizing antibodies rapidly and completely inactivated the wild-type, but not the N370S, acid beta-glucosidase in vitro. Antibodies to human serum albumin and chorionic gonadotropin also developed. The finding of neutralizing antibodies to acid beta-glucosidase during enzyme therapy for Gaucher disease has significant implications for monitoring the therapeutic responses and for potential alternative future therapies for Gaucher disease.

Adolescent↗
PubMed1996

Asp-59 is not important for the catalytic activity of the restriction endonuclease EcoRI.

The amino acid Asp-59 was proposed to be involved in EcoRI catalyzed DNA cleavage (Cheng et al., EMBO J. 13, 3927-35, 1994). We have tested this hypothesis by site directed mutagenesis experiments. The four mutants D59A, D59E, D59G, and D59N bind with similar stability to the specific recognition sequence as wild type EcoRI. The D59E mutant cleaves DNA as fast as the wild type enzyme. Specific activities of the other three mutants are five to tenfold lower. Therefore, we conclude that Asp-59 is not involved in catalysis of the EcoRI restriction endonuclease. Consequences for catalytic mechanisms of EcoRI and other restriction enzymes are discussed.

Amino Acid Sequence↗
PubMed1995

Site-directed mutagenesis in the catalytic center of the restriction endonuclease EcoRI.

The catalytic center of the restriction endonuclease (ENase) EcoRI is structurally homologous to that of EcoRV, BamHI and PvuII. Each of these ENases contains a short motif of three to four amino acid (aa) residues which are positioned in a similar orientation to the scissile phosphodiester bond. We have mutated these aa (Pro90, Asp91, Glu111 and Lys113) in EcoRI to determine their individual roles in catalysis. The replacement of Asp91 and Lys113, respectively, by conservative mutations (Ala91, Asn91, Ala113, Gln113, His113 and Leu113) resulted in a reduction of binding affinity and complete loss of cleavage activity. Only Lys113-->Arg substitution still allows to cleave DNA, albeit with a rate reduced by at least four orders of magnitude. Lys113 seems to stabilize the structure of the wild-type (wt) ENase since all five ENase variants with mutations at this position show a strongly enhanced tendency to aggregate. The Ala and Gln mutants of Glu111 bind the recognition sequence slightly stronger than wt EcoRI and cleave it with a low, but detectable rate. Only the Glu111-->Lys mutant, in which the charge is reversed, shows neither binding nor cleavage activity. Pro90 is not important for catalysis, because the Ala90 mutant cleaves DNA with an only slightly reduced rate. Under star conditions, however, this mutant is even more active than wt EcoRI. Therefore, the charged aa Asp91, Glu111 and Lys113 are essential for catalytic activity of the EcoRI ENase.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1994

Extraosseous extension of Gaucher cell deposits mimicking malignancy.

Two cases are described in which patients with type I Gaucher disease developed extraosseous soft tissue masses consisting of Gaucher cell deposits. In one instance the mass destroyed the posterior cortex of the left distal femur and protruded into the soft tissues. In the second case the lesion involved the proximal tibia and gradually extended into the soft tissues. While the incidence of neoplastic disorder such as lymphoproliferative disease appears to be more common in Gaucher disease patients than in the general population, lesions of benign etiology that mimic these aggressive processes should be considered in the differential diagnosis when cortical destruction with coexisting soft tissue most is found in these patients.

Adult↗
PubMed1993

An in situ study of beta-glucosidase activity in normal and Gaucher fibroblasts with fluorogenic probes.

Beta-glucosidase activity was evaluated in situ by means of fluorogenic probes in normal human fibroblasts and fibroblasts from homozygous carriers of the Gaucher trait. Probe internalization, targeting to lysosomes and post-cleavage probe retention were the primary concerns. Internalization and targeting were attempted by in situ photosensitized labilization of lysosomal membranes, lysosomotropic detergents and the use of low density lipid (LDL) or the receptor ligand apolipoprotein E (ApoE). Post-cleavage increase of fluorescence with fluoresceinyl (bis) beta-glucopyranoside was appreciably above the rather large pre-cleavage emission. In cells incubated overnight with nonylumbelliferyl-beta-glucoside (UG9) in the presence of bovine serum albumin and in the absence of ApoE, the probe was dealt with as a cytotoxic agent, accumulating in a paranuclear cap, most likely comprising elements of the endoplasmic reticulum (ER) and Golgi apparatus. Targeting of UG9 to lysosomes occurred within 1 to 3 h of preincubation in the presence of ApoE. There was some evidence of specificity, as Gaucher fibroblasts exhibited weaker cleavage of UG9 (by 50 per cent or more) compared to normal fibroblasts, but in the Gaucher cells there was some residual beta-glucosidase activity. Cleavage of UG9 was nearly totally suppressed in Gaucher cells treated with the beta-glucosidase inhibitor, conduritol B epoxide, for 24 h to 7 days. Suppression in the control fibroblasts was evident but to a lesser degree. The in situ method of fluorogenic assay established for beta-glucosidase deficiency, is in principle applicable to enzyme deficiencies in other lysosomal storage diseases, or to evaluate enhanced enzyme activity following gene therapy.

Cells, Cultured↗
PubMed1993

MR imaging in adults with Gaucher disease type I: evaluation of marrow involvement and disease activity.

An investigation was conducted to determine the usefulness of magnetic resonance imaging (MRI) in the evaluation of bone marrow involvement in patients with Gaucher disease type I. T1- and T2-weighted images were obtained of the lower extremities of 29 adult patients. Patients were classified into one of three groups based on marrow signal patterns on T1- and T2-weighted images as well as change in signal intensity from T1- to T2-weighted images. An increase in signal intensity from T1- to T2-weighted images was the criterion for an "active process" within the bone marrow. Classification of the 29 patients produced the following results: group A: normal, 4 patients; group B: marrow infiltration, 16 patients; group C: marrow infiltration plus active marrow process, 9 patients. Correlation with clinical findings revealed that all nine patients with evidence of an active marrow process on MRI (group C) had acute bone pain. Conversely, only one of the remaining 20 patients (groups A and B) had bone pain. There was no correlation between disease activity and findings on conventional radiographs. We conclude the MRI provides an excellent noninvasive assessment of the extent and activity of marrow involvement in type I Gaucher disease.

Adult↗
PubMed1990

Mutagenic activity of BKV and JCV in human and other mammalian cells.

We present data suggesting that human polyomaviruses BKV and JCV, widely distributed throughout human populations, are able to induce gene mutations in cultured cells. In this study, using different infecting agents, cell lines to be infected, mutation expression periods, and selection systems, we observed mutagenic effects of varying extent with values of spontaneous mutant frequencies being increased after BKV infection up to 100-fold in BHK cells (6-thioguanine resistance) and nearly 35-fold in virus-transformed human Lesch-Nyhan cells (ouabain resistance). In experiments with BKV the viral mutagenic potential was found to be raised both in moderately uv-irradiated cells, or when wild-type virus was replaced by the variant BKV-IR isolated from a human tumor. Since BKV-IR is defective in the expression of small-t antigen, the viral mutagenicity does not require this protein to be active. BKV was shown to mutate, besides different established cell lines, human peripheral blood lymphocytes. Moreover, as demonstrated by comparing mutagenicities of DNAs from BKV, JCV, and the related polyomavirus SV40, the mutagenic effects of the three viruses do not appear to be essentially different. Implications of these findings are discussed.

Animals↗
PubMed1985

Tartrate-resistant acid phosphatase staining of monocytes in Gaucher disease.

Cytochemical studies were performed on peripheral blood from 30 patients with type 1 Gaucher disease. In 29 of the patients, peripheral blood monocytes stained positively for tartrate-resistant acid phosphatase, whereas monocytes from 18 normal individuals and 14 patients with monocytosis did not. In the Gaucher patients, the percentage of monocyte positivity for tartrate-resistant acid phosphatase ranged from 2 to 97. There was no correlation between the percent monocyte staining and the degree of disease severity, as measured by hepatosplenomegaly, pancytopenia, or extent of bone disease, for the group as a whole. In Gaucher patients who had not undergone splenectomy, however, there was a significant correlation between percent monocyte staining and the degree of hepatosplenomegaly, anemia, and thrombocytopenia. The presence of tartrate-resistant acid phosphatase may be secondary to the lysosomal accumulation of glucosyl ceramide within these monocytes, although this remains to be confirmed. If so, these circulating cells may represent precursors of the Gaucher cells in tissues. Tartrate-resistant acid phosphatase staining of peripheral blood monocytes may be useful as a diagnostic marker for Gaucher type 1 disease and for further studies on the pathogenesis of the disease.

Acid Phosphatase↗
PubMed1979

Defects of bile acid synthesis in Zellweger's syndrome.

Abnormal mitochondrial structure and function have been documented in patients with Zellweger's syndrome (cerebrohepatorenal syndrome). In vitro studies have suggested that the formation of C24 bile acids (chenodeoxycholic acid and cholic acid) from C27 cholesterol requires mitochondrial oxidative clevage of the terminal three carbons of the side chain. Therefore, three patients with Zellweger's syndrome were examined for the presence of mitochondrial defects in bile acid synthesis. All three excreted excessive amounts of 3 alpha, 7 alpha-dihydroxy-5 beta-cholestan-26-oicacid, 3 alpha, 7 alpha, 12 alpha-trihydroxy-5 beta-cholestan-26-oic acid, and 3 alpha, 7 alpha, 12 alpha, 24 xi-tetrahydroxy-5 beta-cholestan-26-oic acid (varanic acid), precursors of chenodeoxycholic acid and cholic acid that have undergone only partial side chain oxidation. These findings give added support to the role of mitochondrial oxidative side chain cleavage in the overall scheme of bile acid synthesis.

Bile Acids and Salts↗
PubMed1979

Enzyme therapy in Fabry disease: differential in vivo plasma clearance and metabolic effectiveness of plasma and splenic alpha-galactosidase A isozymes.

A pilot trial of enzyme replacement with splenic and plasma alpha-galactosidase A (alpha-D-galactosidase; alpha-D-galactoside galactohydrolase, EC 3.2.1.22) isozymes was undertaken in two brothers with Fabry disease, an X-linked glycosphingolipid storage disease. Six unentrapped doses (2000 units/kg) of each isozyme were administered intravenously to the respective recipients during a 117-day period. The circulating half-life of the splenic isozyme was about 10 min, whereas that for the plasma isozyme was approximately 70 min. No immune response was detected by skin and immunodiffusion tests or by alterations in the maximal activity or clearance kinetics for either isozyme after successive administrations. After each dose of the splenic isozyme, the concentration of the accumulated circulating substrate, trihexosylceramide (globotriaosylceramide), decreased maximally (approximately 50% of initial values) in 15 min and returned to preinfusion levels by 2-3 hr. In marked contrast, injection of the plasma isozyme decreased the circulating substrate levels 50-70% by 2-6 hr; the concentrations gradually returned to preinfusion values by 36-72 hr.

Adult↗
PubMedDate not supplied

Nutritional support in patients with systemic scleroderma.

Systemic scleroderma often involves the gastrointestinal tract and can lead to significant malnutrition. Specialized home nutritional support was provided to five patients (one enteral and four parenteral) with restoration of adequate nutritional status, improved quality of life, and few metabolic or technical complications over a period of 12 to 86 months. Three patients eventually died of their disease, one was converted to an oral diet, and one patient remains dependent on total parenteral nutrition. This experience would recommend consideration of home nutritional support in patients with systemic scleroderma when the disease is relatively stable and no major organ failure is present.

Adult↗