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Biomedical subjects

D Linder

Publications and source records attributed to D Linder.

17 recordsLinked to original sources
PubMed1992

Glutamate mutase from Clostridium cochlearium. Purification, cobamide content and stereospecific inhibitors.

Both components, E and S, of the adenosylcobalamin-(coenzyme B12)-dependent glutamate mutase from Clostridium cochlearium were purified. Component S (16 kDa) must be added to component E to obtain activity, although the latter contains substoichiometric amounts of component S besides the major 50-kDa subunit. The enzyme proved to be very similar to that of C. tetanomorphum as described by Barker et al. [Barker, H. A., Rooze, V., Suzuki, F. & Iodice, A. A. (1964) J. Biol. Chem. 239, 3260-3266] but component E of C. cochlearium was more stable and led to the first pure preparation. The pink component E showed a cobamide-like absorbance spectrum with a characteristic maximum at 470 nm indicating the presence of a cob(II)amide, probably Co alpha-[alpha-(aden-9-yl)]-cob(II)amide. A typical cob(II)amide signal at g = 2.23 with hyperfine and superhyperfine splitting was observed by EPR spectroscopy. A cobamide content of about 0.43 mol/mol 50-kDa subunit was determined by cyanolysis. Substitution of the migrating hydrogen at C-4 of glutamate by fluorine yielded the potent competitive inhibitor (2S,4S)-4-fluoroglutamate (Ki = 70 microM). (2R,3RS)-3-Fluoroglutamate (Ki = 600 microM) was also inhibitory. The competitive inhibition by 2-methyleneglutarate (Ki = 400 microM) and (S)-3-methylitaconate (Ki = 100 microM) but not by (RS)-2-methylglutarate suggested the transient formation of an sp2 center during catalysis. However, the presence of an N-terminal pyruvoyl residue was excluded and no evidence for the participation of another electrophilic center in the reaction was obtained.

Amino Acid Isomerases↗
PubMed1992

Isolation and characterization of polyferredoxin from Methanobacterium thermoautotrophicum. The mvhB gene product of the methylviologen-reducing hydrogenase operon.

The methylviologen-reducing hydrogenase operon of Methanobacterium thermoautotrophicum contains an open reading frame, mvhB, the product of which was predicted to have a molecular weight of 44 kDa and to contain as many as 48 iron atoms in 12 [4Fe-4S] clusters, and was therefore suggested to be a polyferredoxin. We have now, for the first time, isolated this polyferredoxin. Its identity with the mvhB gene product was evidenced by a comparison of the N-terminal amino acid sequence. The dark-brown protein of apparent molecular weight 44 kDa was found to contain 53 mol Fe and 43 mol acid-labile sulfur per mol. The UV/visible spectrum showed two maxima at 280 nm and 390 nm, and a shoulder at 308 nm. The A390/A280 ratio was 0.73. The molar extinction coefficient at 390 nm was 170,000 M-1.cm-1. In the dithionite reduced state the protein displayed an EPR spectrum like that of [4Fe-4S] clusters. The results indicate that the mvhB gene product is indeed a polyferredoxin.

Amino Acid Sequence↗
PubMed1992

Chemical composition of Litomosoides carinii microfilarial sheaths.

Litomosoides carinii microfilariae were exsheathed by freezing and thawing, and the sheaths were separated by filtration. Samples of pure sheaths thus obtained were hydrolyzed, methanolyzed or oxidized with nitric acid under pressure at 300 degrees C, respectively, and were analyzed for amino acids, sugars, fatty acids or for metal ions and phosphorus. Almost 75% of the sheath dry weight could thus be accounted for. Amino acids (55 weight %) were the major constituents, and amongst these glutamine and proline (approximately 11% each). The detection of 2% cysteine/cystine indicated the possible presence of disulfide crosslinks. Besides amino acids, approximately 8% of sugars--roughly equimolar amounts of (N-acetyl)galactosamine and uronic acids--1.5% of monovalent cations (Na+ and K+) and 9.5% of phosphate were detected. No appreciable amounts of fatty acids, neutral sugars, neuraminic acid, or (N-acetyl)glucosamine (i.e. no chitin) were found.

Amino Acids↗
PubMed1979

Kidney tubule basement membrane alterations in type II membranoproliferative glomerulonephritis.

Fourteen kidney biopsy specimens from nine patients with type II membranoproliferative glomerulonephritis (MPGN) were examined by electron microscopy for tubular basement membrane (TBM) alterations. In all biopsies, laminal densities, charateristic for type II MPGN, were present in the glomerular basement membranes. The TBM alterations observed included: 1) the presence of laminal, and/or discrete, and/or aggregated densities; 2)focal thickening; 3) multilamination; and, 4) vesicular structures. Laminal densities occurred in 6 of the 9 cases examined. All biopsies had TBM densities representative of at least one of the three forms. The occurrence of electron densities in or near the TBM in type II MPGN may have diagnostic value. In those biopsies where tissue is insufficient for immunofluorescence microscopy and where glomeruli are not found on electron microscopy, an electron microscopic search for densities associated with TBMs would be warranted. Although TBM-associated densities are not pathognomonic for type II MPGN, the observation of such densities, espically laminal densities, would be useful in complementing light miccrscopic and clinical findings.

Adolescent↗
PubMed1977

1,4-alpha-Glucan phosphorylase form Klebsiella pneumoniae covalently couple on porous glass.

A simplified procedure for the preparation of 1,4-alpha-glucan phosphorylase from Klebsiella pneumoniae is described. An 80-fold purification is achieved in two steps with an overall yield of about 50%. The specific activity of the homogeneous enzyme protein is 17.7 units/mg. Compared with glycogen phosphorylase from rabbit muscle the enzyme from K. pneumoniae shows a markedly higher stability against deforming and chaotropic agents. The 1,4-alpha-glucan phosphorylase was covalently bound to porous glass particles by three different methods. Coupling with glutaraldehyde gave the highest specific activity, i.e., 5.6 units/mg of bound protein or 133 units/g of glass with maltodextrin as substrate. This corresponds to about 30% of the specific activity of the soluble enzyme. With substrates of higher molecular weight, such as glycogen or amylopectin, lower relative activity was observed. The immobilized enzyme preparations showed pH activity profiles which were slightly displaced to higher values and exhibited an increased temperature stability.

Drug Stability↗
PubMed1976

1, 4-alpha-Glucan phosphorylase from Klebsiella pneumoniae purification, subunit structure and amino acid composition.

1. A 1,4-alpha-glucan phosphorylase from Klebsiella pneumoniae has been purified about 80-fold with an over-all yield greater than 35%. The purified enzyme has been shown to be homogeneous by gel electrophoresis at different pH-values, by isoelectric focusing, by dodecylsulfate electrophoresis and by ultracentrifugation. 2. The molecular weight of the native enzyme has been determined to be 180 000 by ultra-centrifugation studies, in good agreement with the value of 189 000 estimated by gel permeation chromatography. 3. The enzyme dissociates in the presence of 0.1% dodecylsulfate or 5 M guanidine hydrochloride into polypeptide chains. The molecular weight of these polypeptide chains has been found to be 88 000 by dodecylsulfate polyacrylamide gel electrophoresis and 99 000 by sedimentation equilibrium studies, indicating that the native enzyme is composed of two polypeptide chains. 4. The enzyme contains pyridoxalphosphate with a stoichiometry of two moles per 180 000 g protein, confirming that the 1,4-alpha-glucan phosphorylase from Klebsiella pneumoniae is a dimeric enzyme. 5. The amino acid composition of the enzyme has been determined, and its correspondence to that of 1,4-alpha-glucan phosphorylases from other sources is discussed. 6. The pI of the enzyme has been shown to be 5.3 and its pH-optimum to be about pH 5.9. The enzyme is stable in the range from pH 5.9 to 10.5.

Amino Acids↗
PubMed1976

Culture of cells from the urine and bladder washings of adults.

Cell cultures were attempted from the urine of 5 adults. Of the 28 cultures which were initiated, 21 resulted in the formation of one or more colonies of epithelial cells. Growth could be maintained up to 2 transfers. Poor colony formation was evident in 2 of 4 samples of a saline irrigation of the bladder. This technique may prove to be useful for short-term studies when venipuncture or biopsy are impractical and when epithelial cells are desirable.

Cells, Cultured↗
PubMed1976

Estimating distances from the centromere by means of benign ovarian teratomas in man.

Under the assumption that benign ovarian teratomas in man arise parthenogenically from a germ cell by suppression of the second meiotic division, the distance of a gene from its centromere can be estimated from the observed proportion of heterozygous teratomas collected from heterozygous hosts. The frequency of heterozygous teratomas of heterozygous hosts is equivalent to the frequency of second division segregation at the gene locus which has been used for centromere-related mapping in fungal genetics for more than 40 years. Mapping functions useful for teratoma-based mapping in man are presented.

Chromosome Mapping↗
PubMed1976

Human centromere mapping using teratoma data.

Under the assumption that benign ovarian teratomas arise parthenogenetically from a germ cell by suppression of the second meiotic division, the proportion gamma of heteratomas collected from heterozygous hosts is a measure for the distance between the corresponding gene and its centromere. For proportions gamma less than or equal to 0.3, the mapping function x = gamma/2 applies, where x is the map distance in Morgans.

Cells, Cultured↗
PubMed1975

Parthenogenic origin of benign ovarian teratomas.

To determine the origin of benign cystic teratomas of the ovary, chromosome-banding studies were done on normal tissues and teratomas from five patients. The normal tissues were heterozygous (+/-) for 17 chromosome polymorphisms at or near the centromere, whereas the teratomas were uniformly homozygous (+/+ or -/-). These findings and those employing electrophoretic variants indicate that ovarian teratomas are parthenogenic tumors that arise from a single germ cell after the first meiotic division.

Aminohydrolases↗