[The development and growth of experimental surgery].
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Animals↗
Biomedical subjects
Z Wu
Publications and source records attributed to Z Wu.
[Physicochemical identification of Chinese herbal drugs and materia medica in past 10 years].
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Animals↗
[Study on chemical pattern recognition of Chinese herbal drugs and materia medica in past 10 years].
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Amomum↗
[Indirect determination of industrial chlordane by ultraviolet spectrometry].
Toxicity in chlordane is in proportion to the content of hexachlorocyclopentadiane. Choosing acetone as the solvent, we determined that content by ultraviolet spectrometry, and then calculated roughly the content of this industrial chlordane. The method is cheaper, more rapid than gas chromatography, and suited to rapid testing industrial chlordane.
Chlordan↗
Multiple forms of angiostatin induce apoptosis in endothelial cells.
Angiostatin is a circulating inhibitor of angiogenesis generated by proteolytic cleavage of plasminogen. In this study we have used recombinant human and murine angiostatins (kringles 1-4) as well as native human angiostatin (prepared by elastase digestion of plasminogen [kringles 1-3] or by plasmin autocatalysis in the presence of a free sulfhydryl donor [kringles 1-4]). We report that angiostatin reduces endothelial cell number in a 4-day proliferation assay without affecting cell cycle progression into S-phase (as determined by bromodeoxyuridine labeling). This suggested that the reduction in cell number in the proliferation assay might in part be due to cytotoxicity. This was confirmed by the observation that ethidium homodimer incorporation (a measure of plasma membrane integrity) into endothelial cells was increased by angiostatin in a manner similar to that seen with tumor necrosis factor- (TNF-) and transforming growth factor-beta1 (TGF-beta1), both of which induce apoptosis in endothelial cells. In contrast to TNF- and TGF-beta1, angiostatin did not induce cytotoxicity in human MRC-5 fibroblast, rat smooth muscle, canine MDCK epithelial, or murine B16-F10 melanoma cell lines. Angiostatin-induced apoptosis was confirmed by endothelial cell nuclear acridine orange incorporation as well as by annexin V and TUNEL staining. These in vitro findings point to endothelial cell apoptosis as a mechanism for the antiangiogenic effect of angiostatin in vivo.
Angiostatins↗
Crystal structure of farnesyl protein transferase complexed with a CaaX peptide and farnesyl diphosphate analogue.
The crystallographic structure of acetyl-Cys-Val-Ile-selenoMet-COOH and alpha-hydroxyfarnesylphosphonic acid (alphaHFP) complexed with rat farnesyl protein transferase (FPT) (space group P61, a = b = 174. 13 A, c = 69.71 A, alpha = beta = 90 degrees, gamma = 120 degrees, Rfactor = 21.8%, Rfree = 29.2%, 2.5 A resolution) is reported. In the ternary complex, the bound substrates are within van der Waals contact of each other and the FPT enzyme. alphaHFP binds in an extended conformation in the active-site cavity where positively charged side chains and solvent molecules interact with the phosphate moiety and aromatic side chains pack adjacent to the isoprenoid chain. The backbone of the bound CaaX peptide adopts an extended conformation, and the side chains interact with both FPT and alphaHFP. The cysteine sulfur of the bound peptide coordinates the active-site zinc. Overall, peptide binding and recognition appear to be dominated by side-chain interactions. Comparison of the structures of the ternary complex and unliganded FPT [Park, H., Boduluri, S., Moomaw, J., Casey, P., and Beese, L. (1997) Science 275, 1800-1804] shows that major rearrangements of several active site side chains occur upon substrate binding.
Alkyl and Aryl Transferases↗
JNKK1 organizes a MAP kinase module through specific and sequential interactions with upstream and downstream components mediated by its amino-terminal extension.
MAP kinase (MAPK) cascades are composed of a MAPK, MAPK kinase (MAPKK), and a MAPKK kinase (MAPKKK). Despite the existence of numerous components and ample opportunities for crosstalk, most MAPKs are specifically and distinctly activated. We investigated the basis for specific activation of the JNK subgroup of MAPKs. The specificity of JNK activation is determined by the MAPKK JNKK1, which interacts with the MAPKKK MEKK1 and JNK through its amino-terminal extension. Inactive JNKK1 mutants can disrupt JNK activation by MEKK1 or tumor necrosis factor (TNF) in intact cells only if they contain an intact amino-terminal extension. Mutations in this region interfere with the ability of JNKK1 to respond to TNF but do not affect its activation by physical stressors. As JNK and MEKK1 compete for binding to JNKK1 and activation of JNKK1 prevents its binding to MEKK1, activation of this module is likely to occur through sequential MEKK1:JNKK1 and JNKK1:JNK interactions. These results underscore a role for the amino-terminal extension of MAPKKs in determination of response specificity.
Animals↗
Synthesis and tyrosine kinase inhibitory activity of a series of 2-amino-8H-pyrido[2,3-d]pyrimidines: identification of potent, selective platelet-derived growth factor receptor tyrosine kinase inhibitors.
Screening of a compound library led to the identification of 2-amino-6-(2,6-dichlorophenyl)-8-methylpyrido[2,3-d]pyrimidine (1) as a inhibitor of the platelet-derived growth factor receptor (PDGFr), fibroblast growth factor receptor (FGFr), and c-src tyrosine kinases (TKs). Replacement of the primary amino group at C-2 of 1 with a 4-(N,N-diethylaminoethoxy)phenylamino group yielded 2a, which had greatly increased activity against all three TKs. In the present work, variation of the aromatic group at C-6 and of the alkyl group at N-8 of the pyrido[2,3-d]pyrimidine core provided several analogues that retained potency, including derivatives that were biased toward inhibition of the TK activity of PDGFr. Analogues of 2a with a 3-thiophene or an unsubstituted phenyl group at C-6 were the most potent inhibitors. Compound 54, which had IC50 values of 31, 88, and 31 nM against PDGFr, FGFr, and c-src TK activity, respectively, was active in a variety of PDGF-dependent cellular assays and blocked the in vivo growth of three PDGF-dependent tumor lines.
3T3 Cells↗
Formation of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) by PHA synthase from Ralstonia eutropha.
The acetoacetyl-CoA reductase and the polyhydroxyalkanoate (PHA) synthase from Ralstonia eutropha (formerly Alcaligenes eutrophus) were expressed in Escherichia coli, Klebsiella aerogenes, and PHA-negative mutants of R. eutropha and Pseudomonas putida. While expression in E. coli strains resulted in the accumulation of poly(3-hydroxybutyrate) [PHB], strains of R. eutropha, P. putida and K. aerogenes accumulated poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) [poly(3HB-co-3HHx)] when even chain fatty acids were provided as carbon source, and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [poly(3HB-co-3HV)] when odd chain fatty acids were provided as carbon source. This suggests that fatty acid degradation can be directly accessed employing only the acetoacetyl-CoA reductase and the PHA synthase. This is also the first proof that the PHA synthase from R. eutropha can incorporate 3-hydroxyhexanoate (3HHx) into PHA and has, therefore, a broader substrate specificity than previously described.
3-Hydroxybutyric Acid↗
Replication checkpoint requires phosphorylation of the phosphatase Cdc25 by Cds1 or Chk1.
Checkpoints maintain the order and fidelity of events of the cell cycle by blocking mitosis in response to unreplicated or damaged DNA. In most species this is accomplished by preventing activation of the cell-division kinase Cdc2, which regulates entry into mitosis. The Chk1 kinase, an effector of the DNA-damage checkpoint, phosphorylates Cdc25, an activator of Cdc2. Phosphorylation of Cdc25 promotes its binding to 14-3-3 proteins, preventing it from activating Cdc2. Here we propose that a similar pathway is required for mitotic arrest in the presence of unreplicated DNA (that is, in the replication checkpoint) in fission yeast. We show by mutagenesis that Chk1 functions redundantly with the kinase Cds1 at the replication checkpoint and that both kinases phosphorylate Cdc25 on the same sites, which include serine residues at positions 99, 192 and 359. Mutation of these residues reduces binding of 14-3-3 proteins to Cdc25 in vitro and disrupts the replication checkpoint in vivo. We conclude that both Cds1 and Chk1 regulate the binding of Cdc25 to 14-3-3 proteins as part of the checkpoint response to unreplicated DNA.
14-3-3 Proteins↗
Mutations in domain III alpha of the Mu transposase: evidence suggesting an active site component which interacts with the Mu-host junction.
A series of point mutations was constructed in domain IIIalpha of the Mu A protein. The mutant transposases were purified and assayed for their ability to promote various aspects of the in vitro Mu DNA strand transfer reaction. All mutants with discernable phenotypes were inhibited in stable synapsis (Type 0 or Type 1 complex formation). In contrast, these mutant proteins were capable of LER formation (a transient early reaction intermediate in which the Mu left and right ends have been synapsed with the enhancer), at levels comparable to wild-type transposase. These proteins therefore comprise a novel class of transposase mutants, which are specifically inhibited in stable transpososome assembly. The defect in these proteins was also uniformly suppressed by either Mn2+, or the Mu B protein in the presence of ATP and target DNA. Striking phenotypic similarities were recognized between the domain IIIalpha transposase mutant characteristics noted above, and those for substrate mutants carrying a terminal base-pair substitution at the point of cleavage on the donor molecule. This phenotypic congruence suggests that the alterations in either protein or DNA are exerting an effect on the same step of the reaction i.e., engagement of the terminal nucleotide by the active site. We suggest that domain IIIalpha of the transposase comprises the substrate binding pocket of the active site which interacts with the Mu-host junction.
Amino Acid Sequence↗
Human beta-defensin 2 is a salt-sensitive peptide antibiotic expressed in human lung.
Previous studies have implicated the novel peptide antibiotic human beta-defensin 1 (hBD-1) in the pathogenesis of cystic fibrosis. We describe in this report the isolation and characterization of the second member of this defensin family, human beta-defensin 2 (hBD-2). A cDNA for hBD-2 was identified by homology to hBD-1. hBD-2 is expressed diffusely throughout epithelia of many organs, including the lung, where it is found in the surface epithelia and serous cells of the submucosal glands. A specific antibody made of recombinant peptide detected hBD-2 in airway surface fluid of human lung. The fully processed peptide has broad antibacterial activity against many organisms, which is salt sensitive and synergistic with lysozyme and lactoferrin. These data suggest the existence of a family of beta-defensin molecules on mucosal surfaces that in the aggregate contributes to normal host defense.
Amino Acid Sequence↗
Defining the structural and functional roles of the carboxyl region of the bacteriophage lambda excisionase (Xis) protein.
The bacteriophage lambda excisionase (Xis) protein is required for excisive site-specific recombination. Xis is composed of 72 amino acids and binds cooperatively to two DNA sites (X1 and X2) that are arranged as direct repeats. Alternatively, Xis binds cooperatively with the host-encoded factor for inversion stimulation (FIS) protein at the X1 and F sites, respectively. Here we analyzed the effects of missense substitutions from codon 57 to the carboxyl end of the protein and nonsense mutations that truncate the protein at various positions from residues 60 to 69. We find that all of the mutant proteins promote excision to some extent and interact cooperatively with FIS. Some mutants have no detectible phenotype while others are altered in their abilities to promote excision or to interact cooperatively with integrase (Int). Computer modeling predicts that amino acids from residues 59 to 65 are in an alpha-helix conformation. Mutants with substitutions on one side of the helix at residues 57, 60, 63 and 64 as well as truncated mutants containing 60, 61 or 63 amino acids, fail to interact cooperatively with Int suggesting that this region of the protein forms the interface with Int. Mutants with substitutions at other positions in the putative helix have no detectible phenotype. Residues 66 to 68 may form a reverse turn and the last four amino acids (69 to 72) may not be crucial for the structure or function of the protein.
Amino Acid Sequence↗
2-Substituted aminopyrido[2,3-d]pyrimidin-7(8H)-ones. structure-activity relationships against selected tyrosine kinases and in vitro and in vivo anticancer activity.
While engaged in therapeutic intervention against a number of proliferative diseases, we have discovered the 2-aminopyrido[2, 3-d]pyrimidin-7(8H)-ones as a novel class of potent, broadly active tyrosine kinase (TK) inhibitors. An efficient route was developed that enabled the synthesis of a wide variety of analogues with substitution on several positions of the template. From the lead structure 2, a series of analogues bearing variable substituents at the C-2 position and methyl or ethyl at N-8 was made. Compounds of this series were competitive with ATP and displayed submicromolar to low nanomolar potency against a panel of TKs, including receptor (platelet-derived growth factor, PDGFr; fibroblast growth factor, FGFr; epidermal growth factor, EGFr) and nonreceptor (c-Src) classes. One of the more thoroughly evaluated members was 63 with IC50 values of 0.079 microM (PDGFr), 0.043 microM (bFGFr), 0.044 microM (EGFr), and 0.009 microM (c-Src). In cellular studies, 63 inhibited PDGF-mediated receptor autophosphorylation in a number of cell lines at IC50 values of 0.026-0.002 microM and proliferation of two PDGF-dependent lines at 0.3 microM. It also caused inhibition of soft agar colony formation in three cell lines that overexpress the c-Src TK, with IC50 values of 0.33-1.8 microM. In in vivo studies against a panel of seven xenograft tumor models with known and/or inferred dependence on the EGFr, PDGFr, and c-Src TKs, compound 63 produced a tumor growth delay of 10.6 days against the relatively refractory SK-OV-3 ovarian xenograft and also displayed activity against the HT-29 tumor. In rat oral bioavailability studies, compound 63 plasma concentrations declined in a biexponential manner, and systemic plasma clearance was high relative to liver blood flow. Finally, in rat metabolism studies, HPLC chromatography identified two metabolites of 63, which were proved by mass spectrometry and synthesis to be the primary amine (58) and N-oxide (66). Because of the excellent potency of 63 against selected TKs, in vitro and in vivo studies are underway for this compound in additional tumor models dependent upon PDGFr, FGFr, and c-Src to assess its potential for advancement to clinical trials.
3T3 Cells↗
Characterization of authentic recombinant pea-seed lipoxygenases with distinct properties and reaction mechanisms.
The two major isoforms of lipoxygenase (LOX-2 and LOX-3) from pea (Pisum sativum L. cv. Birte) seeds have been cloned and expressed from full-length cDNAs as soluble, active, non-fusion proteins in Escherichia coli. A comparison of both isoforms purified to apparent homogeneity from E. coli and pea seeds has confirmed the authenticity of the recombinant products and established the properties of the native enzymes. Despite 86% similarity at the amino acid sequence level, the enzymes have distinct properties. They have been characterized in terms of specific activity, Fe content, optimum pH, substrate and product specificity, apparent Km and Vmax for the preferred substrate, linoleic acid, and interfacial behaviour with linoleic acid. We have used this evidence, in addition to EPR spectroscopy of the hydroperoxide-activated enzymes and estimates of kcat/Km, to propose different reaction mechanisms for linoleic acid oxidation for the two isoforms. The differences relate primarily to carbonyl production from linoleic acid for which we propose a mechanism. This implicates the release of a peroxyl radical in an aerobic hydroperoxidase reaction, as the source of the carbonyl compounds formed by dismutation of the liberated peroxyl radical.
Catalysis↗
Stabilization of interleukin-2 mRNA by the c-Jun NH2-terminal kinase pathway.
Signaling pathways that stabilize interleukin-2 (IL-2) messenger RNA (mRNA) in activated T cells were examined. IL-2 mRNA contains at least two cis elements that mediated its stabilization in response to different signals, including activation of c-Jun amino-terminal kinase (JNK). This response was mediated through a cis element encompassing the 5' untranslated region (UTR) and the beginning of the coding region. IL-2 transcripts lacking this 5' element no longer responded to JNK activation but were still responsive to other signals generated during T cell activation, which were probably sensed through the 3' UTR. Thus, multiple elements within IL-2 mRNA modulate its stability in a combinatorial manner, and the JNK pathway controls turnover as well as synthesis of IL-2 mRNA.
CD28 Antigens↗
Absorbance detection of amino acids by laser wave mixing in microbore liquid chromatography.
Nonlinear optical phase conjugation by degenerate four-wave mixing is demonstrated as a sensitive "absorbance" detection method for microbore high-performance liquid chromatography. An argon ion laser operating at the 488-nm line is used as the excitation light source to generate the wave-mixing signal for dabsyl-labeled amino acids. Advantages of the nonlinear laser detection method include: virtually 100% optical signal collection efficiency, generation of the signal in the form of a coherent laser beam, signal measurement against a virtually dark background, reliable detection of small absorbance values, excellent detection sensitivity for both fluorescing and non-fluorescing analytes, relatively simple one-color one-laser optical setup, and low power or energy requirements for continuous-wave or pulsed lasers. Using our one-laser one-color nonlinear laser detector for "absorbance" measurements in liquid chromatography, we report a crude preliminary "injected" detection limit of 780 fmol for glycine.
Algorithms↗
Improvement of beamline 4B9A and results of an X-ray diffraction experiment.
4B9A is a focusing and monochromatic photon beam at the BSRF, which was constructed in 1990. During the second phase of the BSRF program, the surface of the cylindrical mirror has been coated with Pt, covering the original Ni, and the monochromator has been upgraded. The maximum photon energy extends to 11 keV and the intensity has increased about tenfold with respect to the previous intensity at 6 keV. Synchrotron X-ray diffraction patterns for the Hg-1223 (HgBa(2)Ca(2)Cu(3)O(8+delta)) superconducting bulk and thin film have been measured at 1.54014 A. Results indicate that the bulk and film can be indexed as possessing tetragonal symmetry; lattice parameters a = 3.856 A and c = 15.851 A for the bulk Hg-1223 compound, and a = 3.8517 A and c = 15.8511 A for the film. Their structures are similar.
Journal Article↗