Biomedical subjects

Ligong Chen

Publications and source records attributed to Ligong Chen.

8 recordsLinked to original sources

PubMed2006

Spontaneous mobility of GABAA receptor M2 extracellular half relative to noncompetitive antagonist action.

The gamma-aminobutyric acid type A receptor beta(3) homopentamer is spontaneously open and highly sensitive to many noncompetitive antagonists(NCAs) and Zn(2+). Our earlier study of the M2 cytoplasmic half (-1' to 10') established a model in which NCAs bind at pore-lining residues Ala(2)', Thr(6)', and Leu(9)'. To further define transmembrane 2 (M2) structure relative to NCA action, we extended the Cys scanning to the extra cellular half of the beta(3) homopentamer (11' to 20'). Spontaneous disulfides formed with T13'C, L18'C, and E20'C from M2/M2 cross-linking and with I14'C (weak), H17'C, and R19'Con bridging M2/M3 intersubunits, based on single (M2 Cys only) and dual (M2 Cys plus M3 C289S) mutations. Induced disulfides also formed with T16'C, but there were few or none with M11'C, T12'C, and N15'C. These findings show conformational flexibility/mobility in the M2 extracellular half 17' to 20' region interpreted as a deformed beta-like conformation in the open channel. The NCA radioligands used were [(3)H]1-(4-ethynylphenyl)-4-n-propyl-2,6,7-trioxabicyclo[2.2.2]octane ([(3)H]EBOB) and [(3)H]3,3-bis-trifluoromethylbicyclo[2.2.1]heptane-2,2-dicarbonitrile with essentially the same results. NCA binding was disrupted by individual Cys substitutions at 13',14',16',17', and 19'. The inactivity of T13'C/T13'S may have been due to disturbance of the channel gate; I14'S and T16'S showed much better binding activity than their Cys counterparts, and the low activities of H17'C and R19'C were reversed by dithiothreitol. Zn(2+) potency for inhibition of [(3)H]EBOB binding was lowered 346-fold by the mutation H17'A. We propose that NCAs enter their binding site both directly, through the channel pore, and indirectly, through the water cavity of adjacent subunits.

Amino Acid Sequence↗

PubMed2006

Structural model for gamma-aminobutyric acid receptor noncompetitive antagonist binding: widely diverse structures fit the same site.

Several major insecticides, including alpha-endosulfan, lindane, and fipronil, and the botanical picrotoxinin are noncompetitive antagonists (NCAs) for the GABA receptor. We showed earlier that human beta(3) homopentameric GABA(A) receptor recognizes all of the important GABAergic insecticides and reproduces the high insecticide sensitivity and structure-activity relationships of the native insect receptor. Despite large structural diversity, the NCAs are proposed to fit a single binding site in the chloride channel lumen lined by five transmembrane 2 segments. This hypothesis is examined with the beta(3) homopentamer by mutagenesis, pore structure studies, NCA binding, and molecular modeling. The 15 amino acids in the cytoplasmic half of the pore were mutated to cysteine, serine, or other residue for 22 mutants overall. Localization of A-1'C, A2'C, T6'C, and L9'C (index numbers for the transmembrane 2 region) in the channel lumen was established by disulfide cross-linking. Binding of two NCA radioligands [(3)H]1-(4-ethynylphenyl)-4-n-propyl-2,6,7-trioxabicyclo[2.2.2]octane and [(3)H] 3,3-bis-trifluoromethyl-bicyclo[2,2,1]heptane-2,2-dicarbonitrile was dramatically reduced with 8 of the 15 mutated positions, focusing attention on A2', T6', and L9' as proposed binding sites, consistent with earlier mutagenesis studies. The cytoplasmic half of the beta3 homopentamer pore was modeled as an alpha-helix. The six NCAs listed above plus t-butylbicyclophosphorothionate fit the 2' to 9' pore region forming hydrogen bonds with the T6' hydroxyl and hydrophobic interactions with A2', T6', and L9' alkyl substituents, thereby blocking the channel. Thus, widely diverse NCA structures fit the same GABA receptor beta subunit site with important implications for insecticide cross-resistance and selective toxicity between insects and mammals.

Amino Acid Sequence↗

PubMed2004

Syndecan-2 regulates transforming growth factor-beta signaling.

Transforming growth factor-beta (TGF-beta) has multiple functions including increasing extracellular matrix deposition in fibrosis. It functions through a complex family of cell surface receptors that mediate downstream signaling. We report here that a transmembrane heparan sulfate proteoglycan, syndecan-2 (S2), can regulate TGF-beta signaling. S2 protein increased in the renal interstitium in diabetes and regulated TGF-beta-mediated increased matrix deposition in vitro. Transfection of renal papillary fibroblasts with S2 or a S2 construct that has a truncated cytoplasmic domain (S2DeltaS) promoted TGF-beta binding and S2 core protein ectodomain directly bound TGF-beta. Transfection with S2 increased the amounts of type I and type II TGF-beta receptors (TbetaRI and TbetaRII), whereas S2DeltaS was much less effective. In contrast, S2DeltaS dramatically increased the level of type III TGF-beta receptor (TbetaRIII), betaglycan, whereas S2 resulted in a decrease. Syndecan-2 specifically co-immunoprecipitated with betaglycan but not with TbetaRI or TbetaRII. This is a novel mechanism of control of TGF-beta action that may be important in fibrosis.

Binding Sites↗

PubMed2004

Newcastle disease in geese: natural occurrence and experimental infection.

A novel disease entity in domestic geese with unusual epidemiology and pathological changes has emerged in some regions of China since the late 1990s. Velogenic Newcastle disease (ND) viruses were isolated from visceral organs and cloacal swabs of diseased birds in the field. Gross lesions of diseased geese were characterized by extensive necrotic foci in intestinal mucosa, spleen and pancreas. Histological examination revealed multisystemic distribution of lesions. Similar disease was reproduced experimentally in geese not only with two field isolates of goose origin, but also with ND viruses belonging to genotypes VI, VII, VIII and IX. These led us to define the novel disease as clinical ND in geese. Additionally, the goose-originated ND virus isolates could be transmitted from geese to specific pathogen free chickens, illustrating the potential threat they posed to the chicken industry. This is the first detailed report of the natural outbreaks and experimental reproduction of ND in geese, providing evidence that geese could play an important role in the epidemiology of ND.

Animals↗

PubMed2003

Packing structure and self-heating in capillary electrochromatography.

The origin of bubble formation during operation of capillary electrochromatography (CEC) has been an issue of debate. Ohmic heating resulted from current passed through a packed column was proposed as the primary cause. However, this explanation has been questioned on the ground that the current measured in CEC is much lower than that measured with open-tubular separation systems where no bubble formation occurs. To resolve this issue, we carried out a theoretical study correlating self-heating of the electrolyte with packing structure of the column. We used a bundle of capillary tubes, a bundle of two types of capillary tubes and two bundles of capillary tubes connected serially to model, respectively, the flow channels in the column of non-porous particles, in the column of porous particles and in the column of various packing densities. The results from this study indicate that, for columns of homogeneous packing density, the heat output is indeed smaller than that in open-tubular columns of the same dimensions. In this case, the self-heating cannot be a key factor responsible for the bubble formation in CEC. However, for columns of heterogeneous packing density, a large excess of heat release may be produced in column sections of high packing density and, in turn, over-heating in such sections may become the primary cause for the formation of bubbles. It follows from this study that preparation of columns of homogeneous packing structure is essential to obtain reproducible and bubble-free CEC systems.

Chromatography, Micellar Electrokinetic Capillary↗

PubMed2002

Measurement of packing tortuosity and porosity in capillary electrochromatography.

The application of conductivity measurements for packing structure characterization has been extended to a column consisting of a packed section and an open section as typically used in capillary electrochromatography (CEC). Because of the difference in electric conductivity between the packed and open sections, the electric fields applied across the two sections vary, depending on the length of the packed section relative to that of the total column. On the basis of mass conservation law, it can be shown that the ratio of the electric current measured in such a duplex column to that without packing is a function of the length and the geometric structure of the packing bed. Thus, knowing the lengths of the packed section and the whole column, we can readily calculate the obstructive factors, such as the porosity and the tortuosity factor, from the measured conductivity ratio. An example is given to demonstrate the application of this method, with experimental data taken from published work.

Journal Article↗

PubMed2002

Molecular characterization of chicken syndecan-2 proteoglycan.

A partial syndecan-2 sequence (147 bp) was obtained from chicken embryonic fibroblast poly(A)+ RNA by reverse transcription-PCR. This partial sequence was used to produce a 5'-end-labelled probe. A chicken liver cDNA library was screened with this probe, and overlapping clones were obtained encompassing the entire cDNA of 3 kb. The open reading frame encodes a protein of 201 amino acids. The cytoplasmic domain is identical with that of mammalian syndecan-2, and highly similar to those of Xenopus laevis and zebrafish syndecan-2. The transmembrane domain is identical with that of mammalian and zebrafish syndecan-2, and highly similar to that of Xenopus laevis syndecan-2. The ectodomain is 45-62% identical with that of zebrafish, Xenopus laevis and mammalian syndecan-2. Two coding single nucleotide polymorphisms were observed. In vitro transcription and translation yielded a product of 30 kDa. Western blotting of chicken embryonic fibroblast cell lysates with species-specific monoclonal antibody mAb 8.1 showed that chicken syndecan-2 is substituted with heparan sulphate, and that the major form of chicken syndecan-2 isolated from chicken fibroblasts is consistent with the formation of SDS-resistant dimers, which is common for syndecans. A 5'-end-labelled probe hybridized to two mRNA species in chicken embryonic fibroblasts, while Northern analysis with poly(A)+ RNAs from different tissues of chicken embryos showed wide and distinct distributions of chicken syndecan-2 during embryonic development. This pattern was different from that reported for syndecan-4, but consistent with the roles of syndecan-2 in neural maturation and in mesenchymal-matrix interactions.

Amino Acid Sequence↗

PubMedDate not supplied

Synthesis of novel porous magnetic silica microspheres as adsorbents for isolation of genomic DNA.

An improved procedure is described for preparation of novel mesoporous microspheres consisting of magnetic nanoparticles homogeneously dispersed in a silica matrix. The method is based on a three-step process, involving (i) formation of hematite/silica composite microspheres by urea-formaldehyde polymerization, (ii) calcination of the composite particles to remove the organic constituents, and (iii) in situ transformation of the iron oxide in the composites by hydrogen reductive reaction. The as-synthesized magnetite/silica composite microspheres were nearly monodisperse, mesoporous, and magnetizable, with as typical values an average diameter of 3.5 microm, a surface area of 250 m(2)/g, a pore size of 6.03 nm, and a saturation magnetization of 9.82 emu/g. These magnetic particles were tested as adsorbents for isolation of genomic DNA from Saccharomyces cerevisiae cells and maize kernels. The results are quite encouraging as the magnetic particle based protocols lead to the extraction of genomic DNA with satisfactory integrity, yield, and purity. Being hydrophilic in nature, the porous magnetic silica microspheres are considered a good alternative to polystyrene-based magnetic particles for use in biomedical applications where nonspecific adsorption of biomolecules is to be minimized.

Adsorption↗