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Biomedical subjects

M Mathieu

Publications and source records attributed to M Mathieu.

At least 109 records · Page 6Linked to original sources
PubMed1997

Protein engineering with monomeric triosephosphate isomerase (monoTIM): the modelling and structure verification of a seven-residue loop.

Protein engineering experiments have been carried out with loop-1 of monomeric triosephosphate isomerase (monoTIM). Loop-1 of monoTIM is disordered in every crystal structure of liganded monoTIM, but in the wild-type TIM it is a very rigid dimer interface loop. This loop connects the first beta-strand with the first alpha-helix of the TIM-barrel scaffold. The first residue of this loop, Lys13, is a conserved catalytic residue. The protein design studies with loop-1 were aimed at rigidifying this loop such that the Lys13 side chain points in the same direction as seen in wild type. The modelling suggested that the loop should be made one residue shorter. With the modelling package ICM the optimal sequence of a new seven-residue loop-1 was determined and its structure was predicted. The new variant could be expressed and purified and has been characterized. The catalytic activity and stability are very similar to those of monoTIM. The crystal structure (at 2.6 A resolution) shows that the experimental loop-1 structure agrees well with the modelled loop-1 structure. The direct superposition of the seven loop residues of the modelled and experimental structures results in an r.m.s. difference of 0.5 A for the 28 main chain atoms. The good agreement between the predicted structure and the crystal structure shows that the described modelling protocol can be used successfully for the reliable prediction of loop structures.

Amino Acid Sequence↗
PubMed1997

Collaborative study of mosaic tetrasomy 12p or Pallister-Killian syndrome (nineteen fetuses or children).

The difficulties in the diagnosis of Pallister-Killian syndrome are illustrated in this study of nineteen fetuses and children. Diagnosis based on clinical appearance alone is often difficult due to the broad spectrum of clinical anomalies not specific to this syndrome. Due to mosaicism, it is altogether necessary to examine several tissues for the presence of tetrasomy 12p, including circulating lymphocytes in which mosaicism can be as low as 1-3%, amniocytes, chorionic cells and skin fibro-blasts in which mosaicism ranges from 6-100%. When highly suspected on ultrasound examination, the diagnosis recommends prenatal cytogenetic studies because survivors are severely mentally retarded. All the cases are sporadic with only a single preliminary report of recurrence. The cytogenetic diagnosis is therefore helpful in order to reassure family members in regard to genetic counseling.

Abnormalities, Multiple↗
PubMed1997

Gene transfer to human rheumatoid synovial tissue engrafted in SCID mice.

OBJECTIVE: To assess the feasibility of gene therapy in rheumatoid arthritis (RA) and determine the appropriate vector. METHODS: Human rheumatoid synovial tissue from 6 patients with RA was transduced ex vivo with a recombinant retroviral vector (pMFG.nlsLacZ) containing the Escherichia coli beta-galactosidase (beta-gal) gene in a coculture assay in the presence of 20 ng/ml tumor necrosis factor alpha (TNF-alpha) for promoting cell division. We also conducted in vitro infection experiments using an adenoviral vector (AdCMVSpl.LacZ) containing the beta-gal gene. After gene transduction, the synovial tissue was engrafted subcutaneously in 8-week-old severe combined immunodeficiency (SCID) CB17 mice. Beta-gal expression was then monitored as a function of time (up to 21 days) and of virus dose [up to 50 colony forming units (cfu)/cell]. The efficacy of direct in vivo gene transfer was also tested by injection of 10(6) cfu of pMFG.nlsLacZ into rheumatoid synovial tissue engrafted in SCID mice. RESULTS: When recombinant retroviral vector was used, 30 +/- 5% of ex vivo infected synovial cells were positive for staining. In synovial tissue implanted in SCID mice, beta-gal expression declined to 5% after one week, but persisted for at least 21 days. Direct injection of pMFG.nlsLacZ vector into the rheumatoid synovial tissue implanted in SCID mice allowed efficient and stable in vivo infection of the synovial tissue. Ex vivo gene transfer with adenoviral vector resulted in a 98% infection rate of the synovial lining cells. However, beta-gal activity declined 7 days after subcutaneous implantation. CONCLUSION: Highly efficient gene transfer in rheumatoid synovial tissue is achievable with both adenoviral and retroviral vectors, but the results were transient. Exogenous gene transfer through retroviral vectors required stimulation with TNF-alpha for synovial cell division and proviral integration. Direct in vivo gene transfer with recombinant retrovirus was shown to be efficient. Transduction of human synovial tissue engrafted in SCID mice is a potent tool for developing preclinical models of gene therapy in RA.

Adenoviridae↗
PubMed1996

Cloning and characterization of mouse ACF7, a novel member of the dystonin subfamily of actin binding proteins.

We have recently cloned the gene responsible for the mouse neurological disorder dystonia musculorum. The predicted product of this gene, dystonin (Dst), is a neural isoform of bullous pemphigoid antigen 1 (Bpag1) with an N-terminal actin binding domain. Here we report on the cloning and characterization of mouse ACF7. Sequence analysis revealed extended homology of mACF7 with both the actin binding domain (ABD) and the Bpag1 portions of dystonin. Moreover, mACF7 and Dst display similar isoform diversity and encode similar sized transcripts in the nervous system. Phylogenetic analysis of mACF7 and dystonin ABD sequences suggests a recent evolutionary origin and that these proteins form a separate novel subfamily within the beta-spectrin superfamily of actin binding proteins. Given the implication of several actin binding proteins in genetic disorders, it is important to know the pattern of mACF7 expression. mACF7 transcripts are detected principally in lung, brain, spinal cord, skeletal and cardiac muscle, and skin. Intriguingly, mACF7 expression in lung is strongly induced just before birth and is restricted to type II alveolar cells. To determine whether spontaneous mutants that may be defective in mACF7 exist, we have mapped the mACF7 gene to mouse chromosome 4.

Amino Acid Sequence↗
PubMed1996

Crystal structure of enoyl-coenzyme A (CoA) hydratase at 2.5 angstroms resolution: a spiral fold defines the CoA-binding pocket.

The crystal structure of rat liver mitochondrial enoyl-coenzyme A (CoA) hydratase complexed with the potent inhibitor acetoacetyl-CoA has been refined at 2.5 angstroms resolution. This enzyme catalyses the reversible addition of water to alpha,beta-unsaturated enoyl-CoA thioesters, with nearly diffusion-controlled reaction rates for the best substrates. Enoyl-CoA hydratase is a hexamer of six identical subunits of 161 kDa molecular mass for the complex. The hexamer is a dimer of trimers. The monomer is folded into a right-handed spiral of four turns, followed by two small domains which are involved in trimerization. Each turn of the spiral consists of two beta-strands and an alpha-helix. The mechanism for the hydratase/dehydratase reaction follows a syn-stereochemistry, a preference that is opposite to the nonenzymatic reaction. The active-site architecture agrees with this stereochemistry. It confirms the importance of Glu164 as the catalytic acid for providing the alpha-proton during the hydratase reaction. It also shows the importance of Glu144 as the catalytic base for the activation of a water molecule in the hydratase reaction. The comparison of an unliganded and a liganded active site within the same crystal form shows a water molecule in the unliganded subunit. This water molecule is bound between the two catalytic glutamates and could serve as the activated water during catalysis.

Acyl Coenzyme A↗
PubMed1996

Fragile X mutation and FG syndrome-like phenotype.

We present data on 4 mentally retarded brothers, 2 of whom were dizygotic twins with congenital hypotonia, constipation, head size disproportionately large for length or height, and a combination of minor anomalies suggestive of FG syndrome. These brothers have a mentally retarded full sister with similar minor anomalies and an older half-brother with the Martin-Bell syndrome. The mother is mentally retarded; 4 of 7 individuals are positive for fragile X, but all have a CGG expansion ranging from 0.2-2 to 4 kb. Although the phenotype is not completely typical of the FG syndrome and the coincidence of the FMR1 mutation and segregation of the MCA/MR phenotype are highly unlikely, the FMR1 mutation may affect morphogenesis more extensively and differently than the Martin-Bell syndrome does to effect an FG syndromelike phenotype in certain families. This phenotype does not appear to be a contiguous gene syndrome, but an effect of the FMR1 mutation on an adjacent gene must be considered.

Child↗
PubMed1996

Secretion and mitogenic activity of zebrafish FGF3 reveal intermediate properties relative to mouse and Xenopus homologues.

Zebrafish (Brachyodanio rerio) Fgf-3 cDNAs expressed in COS-1 cells give rise to the heterogeneous set of secreted proteins with relative molecular masses in the range of 29-30.5 kDa. These proteins associate strongly with the extracellular matrix but are quantitatively released into the culture medium in the presence of heparin (5 micrograms/ml). Extracellular zebrafish FGF3 (ZFGF3) also contains a smaller sized component that appears to result from an amino-terminal proteolytic cleavage. These properties are similar to those described for Xenopus FGF3 (XFGF3). Receptor binding experiments indicate that ZFGF3 has a higher affinity for the IIIb rather than the IIIc isoform of FGFR2; properties that are more reminiscent of the mouse than the Xenopus homologue. Consistent with the FGF receptor binding properties, ZFGF3 shows a restricted mitogenic potential and a reduced transforming activity on NIH3T3 cells compared to XFGF3. Hybrid proteins made between Xenopus and zebrafish FGF3 implicate the C-terminal region in determining the differences in receptor potential affinities, mitogenic potency and transforming activity. Thus, ZFGF3 shows the structural and secretory properties of XFGF3, but has biological properties more akin to those of the mouse homologue.

Amino Acid Sequence↗
PubMed1996

Molecular cloning of a cDNA encoding the precursor of Ala-Pro-Gly-Trp amide-related neuropeptides from the bivalve mollusc Mytilus edulis.

We isolated and characterised a cDNA clone encoding the precursor of neuropeptides related to the molluscan neuropeptide Ala-Pro-Gly-Trp amide from the marine mussel Mytilus edulis. The preproprotein is posttranslationally processed to generate a 20 amino acid signal peptide together with five sequences of the neuropeptide Arg-Pro-Gly-Trp amide (RPGWamide), one Lys-Pro-Gly-Trp amide (KPGWamide), one Thr-Pro-Gly-Trp amide (TPGW amide) as well as a putative 31 amino acid long C-terminal peptide. In situ hybridisation showed that the gene encoding this precursor is expressed in discrete neurons within the three ganglia of the central nervous system.

Amino Acid Sequence↗
PubMed1996

Prenatal diagnosis of mitochondrial fatty acid oxidation defects.

Amniocytes isolated from two pregnancies at risk for fatty acid oxidation defects were incubated with stable isotopically labelled palmitate, in the presence of L-carnitine, to probe that pathway. The labelled acylcarnitines were then quantitated using tandem mass spectrometry. Amniocytes from a pregnancy at risk for medium-chain acyl-CoA dehydrogenase (MCAD) deficiency produced a characteristic acylcarnitine profile with increased levels of octanoylcarnitine and decanoylcarnitine, indicative of MCAD deficiency. DNA analysis confirmed that the fetus was homozygous for the MCAD A985G mutation. Acylcarnitine and DNA analysis of the infant's blood obtained post-partum confirmed MCAD deficiency. Amniocytes from a pregnancy at risk for an unspecified fat oxidation defect produced increased levels of long-chain acylcarnitines consistent with a deficiency in very-long-chain acyl-CoA dehydrogenase (VLCAD). Measurements of the enzymatic activity confirmed VLCAD deficiency in amniocytes. Acylcarnitine profiles of the infant's blood obtained post-partum in addition to enzyme activities measured in fibroblasts confirmed VLCAD deficiency. The successful prenatal diagnosis of VLCAD and MCAD deficiencies using in vitro probes of fatty acid oxidation in fibroblasts suggests that this approach can potentially recognize many mitochondrial fatty acid oxidation defects even if no prior diagnosis is determined in the family at risk.

Acyl-CoA Dehydrogenase↗
PubMed1996

Stable-isotope selected-ion monitoring quantification of methylmalonic acid in dried filter-paper urine samples.

Urea and creatinine were measured by Kodak Ektachem methods and methylmalonic acid by stable-isotope gas chromatography-mass spectrometry in 24 urine samples from patients with methylmalonic acidaemia. For each sample analyses were performed both directly on the liquid urine and on an aliquot which had been blotted onto filter paper and dried. There was good correlation between the methylmalonate/creatinine ratios in liquid and dried urine (r2 = 0.983). Urine dried on filter paper can be used advantageously for monitoring treatment in patients with this disorder.

Deuterium↗
PubMed1996

Evidence for a fourth locus in Usher syndrome type I.

Usher syndrome type I (US1) is an autosomal recessive condition in which three different genes have been already localised (USH1A, USH1B, and USH1C on chromosomes 14q32, 11q13, and 11p15 respectively). The genetic heterogeneity of US1 has been confirmed in a previous study by linkage analysis of 20 French pedigrees. Here, we report the genetic exclusion of the three previously reported loci in two large multiplex families of Moroccan and Pakistani origin, suggesting the existence of at least a fourth locus in Usher syndrome type I.

Abnormalities, Multiple↗
PubMed1996

Mutation analysis in 24 French patients with glycogen storage disease type 1a.

Both alleles of 24 French glycogen storage disease type 1a patients were sequenced: 14 different mutations allowed the identification of complete genotypes for all the patients. Nine new gene alterations are reported. Five mutations, Q347X, R83C, D38V, G188R, and 158 del C, account for 75% of the mutated alleles. These data show that the molecular pathology of the glucose-6-phosphatase gene is heterogeneous in this population. Complete genotyping of the index case by systematic sequencing is necessary to allow prenatal diagnosis in chorionic villi for at risk couples.

Base Sequence↗
PubMed1996

[Vectors of gene transfer. Applications in pneumology].

Human gene therapy in 1996 is a reality with almost 100 trials world-wide involving nearly 450 patients. The stakes are enormous. To assure the efficacy and safety of such a regime is mainly a problem of vector to carry the chosen nucleotide sequences. The ideal vector should have as a unique function that of the transport of the gene of therapeutic interest to the desired area at the proper quantity. The ideal vector does not currently exist and each system, viral or non-viral has both advantages and inconveniences.

Base Sequence↗
PubMed1996

The role of YAG-laser iridotomy in pigment dispersion syndrome.

We determined iris configuration and posterior-chamber depth before and after YAG-laser iridotomy in 20 eyes with pigment dispersion syndrome using high-resolution ultrasound and measured the intraocular pressure after a mean of 9.1 months. The iris configuration and posterior-chamber depth of our patients were compared with those of an age- and refraction-matched control group of 20 eyes without pigment dispersion. In the patient group, 8 eyes had a concave iris configuration with shallow posterior chambers and 12 eyes had planar irides. In the former group the posterior-chamber depth increased significantly after treatment. In the latter group it did not change. In the normal control group, three subjects also had concave irides. There was a significant correlation between posterior-chamber depth and refraction in patients with pigment dispersion syndrome, but not in the normal control group. After the follow-up period the intraocular pressure of treated eyes was not significantly lower than that of untreated eyes. Iris concavity can also occur in normal individuals and may therefore be only a pathogenetic cofactor in the pigment dispersion syndrome. Laser iridotomy proved to flatten irides but did not significantly decrease the intraocular pressure.

Adult↗
PubMed1995

Receptor binding and mitogenic properties of mouse fibroblast growth factor 3. Modulation of response by heparin.

fgf3 has been implicated in the embryonic and fetal development of the mouse and as an oncogene in murine breast cancer. We describe a procedure to purify the product of the mouse fgf3 gene and show it to be a potent mitogen for some epithelial cell lines. Using a receptor binding competition assay, Fgf3 was shown to bind with high affinity to the IIIb isoforms of Fgf receptor (FgfR) 1 and FgfR2 (ID50 = approximately 0.8 nM) and with a lower affinity to the IIIc variant of FgfR2 (ID50 = approximately 9 nM). No competition for the binding of 125I-Fgf1 was observed for FgfR1 (IIIc), FgfR3 (IIIb and IIIc), or FgfR4. Mitogenicity assays using BaF3 cells containing individual Fgf receptors showed a pattern of response in agreement with the receptor binding results. A comparison of two mammary epithelial cell lines showed a marked difference of potency and dependence upon heparin in their response to mouse Fgf3, suggesting a complex interaction between the ligand and its low and high affinity receptors.

3T3 Cells↗