Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0033036 (APC)
 10,214 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Glycogen synthase kinase 3 (GSK-3), an element of the Wnt signalling pathway, plays a key role in numerous cellular processes including cell proliferation, embryonic development, and neuronal functions. It is directly involved in diseases such as cancer (by controlling apoptosis and the levels of beta-catenin and cyclin D1), Alzheimer's disease (tau hyperphosphorylation), and diabetes (as a downstream element of insulin action, GSK-3 regulates glycogen and lipid synthesis). We describe here a rapid and efficient method for the purification of GSK-3 by affinity chromatography on an immobilized fragment of axin. Axin is a docking protein which interacts with GSK-3ss, beta-catenin, phosphatase 2A, and APC. A polyhistidine-tagged axin peptide (residues 419-672) was produced in Escherichia coli and either immobilized on Ni-NTA agarose beads or purified and immobilized on CNBr-activated Sepharose 4B. These "Axin-His6" matrices were found to selectively bind recombinant rat GSK-3 beta and native GSK-3 from yeast, sea urchin embryos, and porcine brain. The affinity-purified enzymes displayed high kinase activity. This single step purification method provides a convenient tool to follow the status of GSK-3 (protein level, phosphorylation state, kinase activity) under various physiological settings. It also provides a simple and efficient way to purify large amounts of active recombinant or native GSK-3 for screening purposes.
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PMID:Purification of GSK-3 by affinity chromatography on immobilized axin. 1108 79

Previous uranium mining in the "Wismut" region in Germany enhanced environmental distribution of heavy metals and radionuclides. Carryover effects may now lead to contamination of locally produced foods. Compounds of "Wismut" origin are probably genotoxic via their irradiating components (radon) or by interacting directly with cellular macromolecules. To assess possible hazards, we investigated the genotoxic effects of uranyl nitrilotriacetate (U-NTA) in human colon tumor cells (HT29 clone 19A), adenoma cells (LT97), and nontransformed primary colon cells. These are target cells of oral exposure to environmentally contaminated foods and represent different cellular stages during colorectal carcinogenesis. Colon cells were incubated with U-NTA. Cell survival, cytotoxicity, cellular glutathione (GSH) levels, genotoxicity, and DNA repair capacity (comet assay), as well as gene- and chromosome-specific damage combination of comet assay and fluorescence in situ hybridization [FISH], 24-color FISH) were determined. U-NTA inhibited growth of HT29 clone 19A cells (75-2000 microM, 72 h) and increased GSH (125-2000 microM, 24 h). U-NTA was genotoxic (1000 microM, 30 min) but did not inhibit the repair of DNA damage caused by hydrogen peroxide (H(2)O(2)), 4-hydroxynonenal, and 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]-pyridine. U-NTA was also genotoxic in LT97 cells and primary colon cells, where it additionally increased migration of TP53 into the comet tail. In LT97 cells, 0.5-2mM U-NTA increased chromosomal aberrations in chromosomes 5, 12, and 17, which harbor the tumor-related genes APC, KRAS, and TP53. It may be concluded that uranium compounds could increase alimentary genotoxic exposure in humans if they reach the food chain in sufficient amounts.
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PMID:Uranyl nitrilotriacetate, a stabilized salt of uranium, is genotoxic in nontransformed human colon cells and in the human colon adenoma cell line LT97. 1684 May 63

The gene (pac) encoding beta-lactam acylase from Bacillus badius was cloned and expressed in Escherichia coli. The pac gene was identified by polymerase chain reaction (PCR) using degenerated primers, on the basis of conserved amino acid residues. By using single specific primer PCR (SSP-PCR) and direct genome sequencing, a complete pac gene with its promoter region was obtained. The ORF consisted of 2415 bp and the deduced amino acid sequence indicated that the enzyme is synthesized as a preproenzyme with a signal sequence, an alpha-subunit, a spacer peptide and a beta-subunit. The pac gene was expressed with its own promoter in different E. coli host strains and a maximum recombinant PAC (1820 U l(-1)) was obtained in E. coli DH5alpha. The recombinant PAC was purified by Ni-NTA chromatography and the purified PAC had two subunits with apparent molecular masses of 25 and 62 kDa. This enzyme exhibited a high thermostability with a maximum activity at 50 degrees C. This enzyme showed stability over a wide pH range (pH 6.0-8.5) with a maximum activity at pH 7.0 and activity on a wide beta-lactam substrate range. The K(m) values obtained for the hydrolysis of penicillin G and a chromogenic substrate, 6-nitro-3-phenylacetylamidobenzoic acid, from B. badius PAC were 39 and 41 microM, respectively. The PAC activity was competitively inhibited by PAA (K(i), 108 microM) and noncompetitively by 6-APA (K(i), 17 mM). The constitutive production of B. badius PAC in E. coli and its easier purification together with the advantageous properties, such as thermostability, pH stability and broad substrate specificity, make this as a novel enzyme suitable for beta-lactam industry.
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PMID:Molecular cloning and characterization of thermostable beta-lactam acylase with broad substrate specificity from Bacillus badius. 1760 62

PAC1 is a pituitary adenylate cyclase-activating polypeptide (PACAP) preferring receptor, which is abundant in the central and peripheral nervous systems. PAC1 belongs to the class B family of G protein-coupled receptors (GPCRs). The N-terminal first extracellular (EC1) domain of PAC1 is responsible for ligand recognition and binding. In this study, the recombinant EC1 domain of the PAC1 normal (N) form (amino acids 21-155) with 6His tag at the C-terminus (named PAC1-EC1(N)) was first expressed in an Escherichia coli strain and purified by an Ni-NTA affinity column. About 6-8mg of recombinant PAC1-EC1(N) protein with purity above 95% was produced from 1L of bacterial culture. Mass spectrum and western blot were used to identify the recombinant PAC1-EC1(N). Intrinsic tryptophan fluorescence (ITF) assays showed that the purified PAC1-EC1(N) protein was able to recognize and bind to the PAC1 selective agonist maxadilan, the antagonist M65 and vasoactive intestinal polypeptide (VIP). Maxadilan and M65 had higher affinities for PAC1-EC1(N) than VIP. The results of MTT assays showed that PAC1-EC1(N) stimulated the viability of PAC-CHO cells but blocked the effects of maxadilan on the proliferation of CHO cells expressing PAC1 (PAC1-CHO), indicating that the functional soluble PAC1-EC1(N) may act as a regulator for the activation of PAC1.
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PMID:The functional recombinant first extracellular (EC1) domain of PACAP receptor PAC1 normal form (PAC1-EC1(N)) recognizes selective ligands and stimulates the proliferation of PAC1-CHO cells. 2055 98