Biomedical subjects

S W Michnick

Publications and source records attributed to S W Michnick.

24 records · Page 2Linked to original sources

PubMed1991

Proton and nitrogen sequential assignments and secondary structure determination of the human FK506 and rapamycin binding protein.

Sequential 1H and 15N assignments of human FKBP, a cytosolic binding protein for the immunosuppressive agents FK506 and rapamycin, are reported. A combination of homonuclear and relayed heteronuclear experiments has enabled assignment of 98 of 99 backbone amide NHs, 119 of 120 C alpha Hs, 97 of 99 non-proline amide 15Ns, and 375 of 412 side-chain resonances of this 107-residue protein. Long-range NOEs are used to demonstrate that FKBP has a novel folding topology consisting of a five-stranded antiparallel beta sheet with +3, +1, -3, +1 loop connectivity.

Anti-Bacterial Agents↗

PubMed1991

Solution structure of FKBP, a rotamase enzyme and receptor for FK506 and rapamycin.

Immunophilins, when complexed to immunosuppressive ligands, appear to inhibit signal transduction pathways that result in exocytosis and transcription. The solution structure of one of these, the human FK506 and rapamycin binding protein (FKBP), has been determined by nuclear magnetic resonance (NMR). FKBP has a previously unobserved antiparallel beta-sheet folding topology that results in a novel loop crossing and produces a large cavity lined by a conserved array of aromatic residues; this cavity serves as the rotamase active site and drug-binding pocket. There are other significant structural features (such as a protruding positively charged loop and an apparently flexible loop) that may be involved in the biological activity of FKBP.

Anti-Bacterial Agents↗

PubMed1989

Separation of the complex asparagine-linked oligosaccharides of the glycoprotein fetuin and elucidation of three triantennary structures having sialic acids linked only to galactose residues.

Asparagine-linked oligosaccharides of the glycoprotein fetuin were isolated as reducing oligosaccharides after hydrazinolysis/re-N-acetylation/mild acid treatment of the Pronase-digested glycoprotein. The sialylated oligosaccharides were separated by high-performance liquid chromatography in two different systems, which resulted in greater than 35 fractions, comprising di-, tri-, tetra-, and pentasialylated oligosaccharides. The major components were isomeric structures comprising the tri- and tetrasialylated fractions. In this and the accompanying paper (Cumming et al., 1989), the structures of 10 of the major components of the tri-, tetra-, and pentasialylated oligosaccharide fractions are described. Separation protocols and three isolated structures having sialic acid linked only to galactose are presented in this paper.

Asparagine↗

PubMed1989

Structures of asparagine-linked oligosaccharides of the glycoprotein fetuin having sialic acid linked to N-acetylglucosamine.

In the accompanying paper (Bendiak et al., 1989), the separation of a series of oligosaccharides released from asparagine residues of fetuin was described. A series of NMR experiments, which included one- and two-dimensional nuclear Overhauser enhancement, two-dimensional correlation spectroscopy, and two-dimensional relayed-coherence spectroscopy, as well as permethylation analyses, established a Gal beta 1----3(NeuAc alpha 2----6)GlcNAc beta 1----4Man unit common to a series of purified structures. These oligosaccharides contained either three, four, or five glycosidically linked sialic acid residues. The NeuAc residue in alpha 2----6 linkage to GlcNAc gives rise to diagnostic chemical shift perturbations of particular proton signals in the oligosaccharides.

Acetylglucosamine↗

PubMed1989

Control of glycoprotein synthesis. Detection and characterization of a novel branching enzyme from hen oviduct, UDP-N-acetylglucosamine:GlcNAc beta 1-6 (GlcNAc beta 1-2)Man alpha-R (GlcNAc to Man) beta-4-N-acetylglucosaminyltransferase VI.

Hen oviduct membranes were shown to contain high activity of a novel enzyme, UDP-GlcNac:GlcNAc beta 1-6(GlcNAc beta 1-2) Man alpha-R (GlcNAc to Man) beta 4-GlcNAc-transferase VI. The enzyme was shown to transfer GlcNAc in beta 1-4 linkage to the D-mannose residue of GlcNAc beta 1-6 (GlcNAc beta 1-2) Man alpha-R where R is either 1-6Man beta-(CH2)8COOCH3 or methyl. Radioactive enzyme products were purified by several chromatographic steps, including high performance liquid chromatography, and structures were determined by proton nmr, fast atom bombardment-mass spectrometry, and methylation analysis to be GlcNAc beta 1-6 ([14C]GlcNAc beta 1-4) (GlcNAc beta 1-2) Man alpha-R. The enzyme is stimulated by Triton X-100 and has optimum activity at a relatively high MnCl2 concentration of about 100 mM; Co2+, Mg2+, and Ca2+ could partially substitute for Mn2+. A tissue survey demonstrated high GlcNAc-transferase VI activity in hen oviduct and lower activity in chicken liver and colon, duck colon, and turkey intestine. No activity was found in mammalian tissues. Hen oviduct membranes cannot act on GlcNAc beta 1-6Man alpha-R but have a beta 4-GlcNAc-transferase activity that converts GlcNAc beta 1-2Man alpha-R to GlcNAc beta 1-4(GlcNAc beta 1-2) Man alpha-R where R is either 1-6Man beta-(CH2)8COOCH3 or 1-6Man beta methyl. The latter activity is probably due to GlcNAc-transferase IV which preferentially adds GlcNAc in beta 1-4 linkage to the Man alpha 1-3 arm of the GlcNAc beta 1-2Man alpha 1-6(GlcNAc beta 1-2Man alpha 1-3)Man beta 1-4GlcNAc beta 1-4GlcNAc-Asn core structure of asparagine-linked glycans. The minimum structural requirement for a substrate of beta 4-GlcNAc-transferase VI is therefore the trisaccharide GlcNAc beta 1-6(GlcNAc beta 1-2) Man alpha-; this trisaccharide is found on the Man alpha 6 arm of many branched complex asparagine-linked oligosaccharides. The data suggest that GlcNAc-transferase VI acts after the synthesis of the GlcNAc beta 1-2Man alpha 1-3-, GlcNAc beta 1-2Man alpha 1-6-, and GlcNAc beta 1-6 Man alpha 1-6-branches by GlcNAc-transferases I, II, and V, respectively, and is responsible for the synthesis of branched oligosaccharides containing the GlcNAc beta 1-6(GlcNAc beta 1-4)(GlcNAc beta 1-2)Man alpha 1-6Man beta moiety.

Acetylglucosamine↗

PubMed1989

Oligosaccharide-protein interactions: a three-dimensional view.

For carbohydrates to serve as recognition elements in cellular function, there must be 'receptors' which are capable of distinguishing between the multitude of oligosaccharide structures generated by a cell. Generally these receptors are assumed to be proteins, and the plant lectins have been used as model systems to examine the molecular basis for specificity in such interactions. Three aspects of the specificity of oligosaccharide-protein interactions will be discussed: (1) the conformational flexibility of oligosaccharides will be demonstrated through a quantitative analysis of nuclear magnetic resonance measurements; (2) a comparison of the measured and calculated values for the entropy barrier to oligosaccharide binding will be used to argue that the barrier arises from a loss of this conformational flexibility upon binding to the lectin (this conclusion is also supported by X-ray crystallographic studies); and (3) the thermodynamic model can be extended to the binding of glycoproteins to receptors and the high affinity of these interactions explained by either multivalency or fixation of the oligosaccharide in the 'correct' three-dimensional structure through interaction with the protein moiety.

Animals↗