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C Fenselau

Publications and source records attributed to C Fenselau.

At least 91 records · Page 5Linked to original sources
PubMed1988

2,4-Dinitrophenylhydrazides of polysialogangliosides.

Treatment with 2,4-dinitrophenylhydrazine HCl in the presence of dicyclohexylcarbodiimide, converts gangliosides to their dinitrophenylhydrazides. This derivatization is the basis of a useful method for HPLC determination of gangliosides (K. Miyazaki, N. Okamura, Y. Kishimoto and Y. C. Lee (1986) Biochem. J. 235, 755-761). This procedure, however, yields two different GT1b products. By characterizing these two products using plasma desorption mass spectrometry, proton magnetic resonance and other chemical and physical techniques, we found that either one or two of the three sialic acid carboxyl groups in GT1b, were converted to dinitrophenylhydrazides. The remaining underivatized carboxyl groups formed lactones with hydroxyl groups from other carbohydrate residues. Also, while sialic acid residues of GD1a were fully derivatized, only one sialic acid in GD1b, two sialic acids in GT1a and two in GQ1b were converted to dinitrophenylhydrazides, the remaining carboxyl groups probably forming lactones. Sialic acid residues between galactose of the gangliotetraose chain and another sialic acid in polysialogangliosides appear to be underivatized possibly because of steric hindrance.

Chemical Phenomena↗
PubMed1988

Enzymatic mechanisms of resistance to alkylating agents in tumor cells and normal tissues.

The presence in tumor cells and in normal cells of enzymes which metabolize and inactive alkylating agents appears to play a major role in determining the effectiveness of alkylating agents against human tumors and the toxicities of these agents to normal tissues. The enzyme aldehyde dehydrogenase appears to protect bone marrow and the gastrointestinal tract against toxicity from cyclophosphamide and other closely related oxazophosphorine agents. The presence of this enzyme in bone marrow stem cells facilitates the elimination of tumor cells from bone marrow suspensions, with preservation of the ability of the marrow suspension to reconstitute normal hematopoiesis in a patient. A variety of mouse and human tumors has been shown to be resistant to cyclophosphamide on the basis of an elevated aldehyde dehydrogenase content. The clinical significance of this type of resistance is currently being explored. Increased levels of glutathione-S-transferase have been shown to be associated with cellular resistance to a variety of alkylating agents. We have identified and characterized the conjugates of nitrogen mustards with glutathione. The formation of these conjugates is catalyzed by glutathione-S-transferase. The further study and characterization of these specific reactions should contribute to the understanding and quantitation of this type of alkylating agent resistance.

Aldehyde Dehydrogenase↗
PubMed1988

Use of immobilized enzymes in drug metabolism studies.

The immobilization of drug-metabolizing enzymes onto polymeric supports offers several advantages over use of conventional microsomal or soluble enzyme preparations. These include increased storage stability, facilitated separation of products from the incubation mixture, the ability to recover and reuse the enzyme catalyst, and in many cases, stabilization of the tertiary structure of membrane-bound enzymes. Attachment of the protein to the solid support may be accomplished by adsorption, covalent bonding, or entrapment techniques. This methodology has been successfully utilized for studies with such enzymes as cytochrome P-450, UDP-glucuronyltransferases, glutathione S-transferases, S-methyltransferases, and N-acetyltransferases. Although often employed for the synthesis of xenobiotic metabolites, immobilized enzymes have been used for mechanistic and relative reactivity studies, limited kinetic studies, and extracorporeal detoxification. Co-immobilization of multiple drug-metabolizing enzyme systems has made possible the sequential formation of metabolites arising from oxidation followed by conjugation. Immobilized enzymes may also be used in the prediction of species-dependent metabolic pathways. The potential for large-scale synthesis of drug metabolites using this methodology has been explored.

Animals↗
PubMed1987

Phosphorylation heterogeneity of tryptic phosphopeptides of chicken riboflavin-binding protein.

The tryptic phosphopeptide of hen egg white riboflavin-binding protein has been found to exist as a mixture of peptides which differ only with respect to the number of covalently bound phosphoryl groups. Anion-exchange chromatography was used to separate homologues of the tryptic phosphopeptide of egg white riboflavin-binding protein. Four peptide peaks were obtained and analyzed using plasma desorption mass spectrometry. Molecular ions obtained agree closely with calculated molecular weight values for phosphopeptides with 8, 7 and 5 phosphoryl groups. Amino acid analyses showed that the octa- and hepta-phosphorylated peptides were pure and had the same amino acid compositions.

Animals↗
PubMed1987

Non-enzymatic formation of insulin-glutathione mixed disulfides: evidence for a transient species by plasma desorption mass spectrometry.

Formation of insulin-glutathione mixed disulfides takes place under the conditions of 0.1 M ammonium acetate, neutral pH and without the presence of any enzyme. Using a SH-free glutathione-agarose column it is demonstrated that the interaction of insulin with glutathione is specific, and increasing the incubation time between these two peptides results in the reduction of insulin disulfide bonds and the production of A and B chains.

Animals↗
PubMed1987

Mass spectral analysis of complex lipids desorbed directly from lyophilized membranes and cells.

Three desorption ionization techniques--laser desorption, plasma desorption and fast atom bombardment mass spectrometry--have been applied to lyophilized cells, membranes, lysed cells and various extracts. It has been shown that intact polar lipids are selectively desorbed from biological membranes by these methods and that their mass spectra provide "fingerprints" which reflect the unique biochemical composition of each class of cell or membrane.

Animals↗
PubMed1985

Phosphorylation sites in riboflavin-binding protein characterized by fast atom bombardment mass spectrometry.

The capability of fast atom bombardment mass spectrometry for characterization of phosphorylation sites in a tryptic peptide from chicken egg yolk riboflavin-binding protein has been evaluated. The quality of information about molecular weight, amino acid sequence, phosphorylation sites, and microheterogeneity is evaluated as a function of the sign of the ions analyzed, the nature of the counter ions associated with the phosphate substituents, sample matrix, and various instrumental parameters. The intact octaphosphorylated 23-residue peptide was found to be susceptible to mass spectral analysis. Information from the negative ion spectrum was used in conjunction with complete sequence information and experiments which showed that all phosphates were attached to serine residues. Phosphorylated and unphosphorylated serine residues were identified and the sample was shown to be homogeneously octaphosphorylated.

Amino Acid Sequence↗
PubMed1985

Mass spectral analysis of murine epidermal growth factor.

Fast atom bombardment mass spectrometry has been used to characterize epidermal growth factor isolated from mouse submaxillary glands. The preparation is found to consist of two peptides, one of which has the average molecular weight predicted for the familiar gene product. The molecular weight of the second component is found to be reduced by the mass of one asparagine residue. These observations are discussed in light of previous reports of heterogeneity.

Amino Acid Sequence↗
PubMed1983

Position of ester groups in the lipid A backbone of lipopolysaccharides obtained from Salmonella typhimurium.

Lipopolysaccharides extracted from the heptoseless mutant of Salmonella typhimurium G30/C21 were hydrolyzed with either 0.1 N HCl at 100 degrees C or treated twice with 20 mM sodium acetate, pH 4.5, at 100 degrees C for 45 min and finally purified by preparative thin layer chromatography to yield a structural series of mono- and diphosphoryl lipid A, respectively. Positive ion fast atom bombardment mass spectrometry of the diphosphoryl lipid A TLC-3 (a highly acylated major band) showed a major component with (M + H)+ ion of mass 1798, which fragmented to yield a (M - H2PO4)+ ion of mass 1700. Cleavage at the glycosidic bond gave rise to an oxonium ion fragment of mass 1087. In conjunction with other studies, this establishes the molecular formula and Mr of the major component to be C94H178N2O25P2 and 1797.2 (as the free acid), respectively. Similar analysis of monophosphoryl lipid A TLC-3 produced an (M + H)+ peak at m/z 1718, (M + Na)+ adduct peak at m/z 1740, and a fragment of mass 1087. The spectrum of monophosphoryl lipid A TLC-5 was devoid of the m/z 1087 peak and instead contained the phosphorylated oxonium ion of mass 876. This fragment ion is assigned as the distal subunit, and these results show that the distal subunit of the major lipid A TLC-3 contains two hydroxymyristoyl, one myristoyl, and one lauroyl residues, whereas the reducing end subunit contains two hydroxymyristoyl groups. A revised structure of the lipid A backbone in lipopolysaccharides of S. typhimurium is proposed.

Chromatography, Thin Layer↗
PubMed1983

Purification and characterization of the heat-stable factors essential for the conversion of lignoceric acid to cerebronic acid and glutamic acid: identification of N-acetyl-L-aspartic acid.

The conversion of lignoceric acid to cerebronic acid, ceramides, cerebrosides, and glutamic acid is catalyzed by a rat brain particulate preparation. The heat-stable factor, prepared from calf cerebellum, together with the heat-labile factor, a pyridine nucleotide, and Mg2+ are essential to all of these metabolic pathways. Our previous work showed that the heat-stable factor is composed of at least two components, HSF-1 and HSF-2, and identified HSF-2 as D-glucose-6-phosphate. In the current investigation, HSF-1 was further purified and found to be N-acetyl-L-aspartic acid. In addition, it was discovered that a third component, HSF-3, is also required for heat-stable factor activity. A reconstituted system composed of N-acetylaspartic acid, glucose-6-phosphate, and HSF-3 fully replaced the heat-stable factor essential for the conversion of lignoceric acid to cerebronic acid and glutamic acid. The reconstituted heat-stable factor did not show the initial time lag always observed with the crude heat-stable factor.

Animals↗
PubMed1983

Dipalmitoylphosphatidylcholine in amniotic fluid quantified by fast-atom-bombardment mass spectrometry.

Dipalmitoylphosphatidylcholine (DPC) is quantified by taking advantage of stable-isotope-labeled d9-DPC as internal standard. Use of a mass spectrometer to measure the ratio of d9/d0 makes this procedure a quantitative one. d9-DPC was synthesized by refluxing dipalmitoylethanolamine with d3-methyl iodide in methanol in the presence of sodium bicarbonate for 26 h. The yield of d9-phosphatidylcholine (d9-lecithin) was 89% after column-chromatographic purification. Fast atom bombardment was used to desorb the preformed phosphatidylcholine ions in a mass spectrometer of Nier-Johnson geometry. In our assessment of accuracy and precision of this technique, we found a correlation coefficient of 0.9994 between signal and sample concentration. The method was less precise when the total d0- plus d9-DPC was less than 0.2 micrograms or when the ratio of d0- to d9-lecithin exceeded 100. The within-run CV was about 1.0%. The amount of DPC in amniotic fluid samples assessed by mass spectrometry was compared with results for total phosphatidylcholine quantified by thin-layer chromatography. The fate of DPC in various laboratory manipulations was also studied.

Amniotic Fluid↗