Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Plasmids containing the chromosome region of Escherichia coli encoding phoM, whose product is a positive regulator of alkaline phosphatase expression, were isolated from the Clarke and Carbon plasmid bank. A 9.9-kilobase EcoRI fragment of plasmid pLC17-39 (subcloned into pBR322) was able to complement both phoM and thrB mutations. Restriction endonuclease analysis and in vitro mutagenesis of the hybird plasmids enabled the localization of the phoM gene locus to 3 kilobases of the cloned chromosomal fragment. The phoM gene product was identified, with maxicell techniques, as a protein with an approximate molecular weight of 55,000. A phoM-lacZ protein fusion was constructed by using a plasmid carrying the phoM gene and a derivative of phage lambda, lambda plac Mu2. Restriction endonuclease analysis of the plasmid carrying the fusion indicated that phoM is transcribed in a clockwise direction on the circular E. coli chromosome. Analysis of strains bearing the fusion on a multiple-copy plasmid or integrated at the lambda attachment site of the chromosome indicated that the synthesis of the phoM gene product was unaffected by phosphate limitation of growth. The expression of the phoM gene was studied in strains with mutations in genes encoding effectors of the pho regulon. A threefold increase in phoM expression was seen in a phoU strain in comparison with the wild-type strain.
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PMID:Identification of the phoM gene product and its regulation in Escherichia coli K-12. 633 29

Carbon-13 nuclear magnetic resonance (13C NMR) of Escherichia coli alkaline phosphatase labeled biosynthetically with beta,beta-[gamma-13C]dideuteriohistidine has been used to determine the number and identity of the histidine residues that participate in metal ion coordination at the three classes of binding sites in this dimeric Zn2+ metalloenzyme. Detailed 13C NMR titrations of the apoenzyme with 113Cd2+ and Mg2+, in conjunction with parallel 13 Cd NMR measurements [Otvos, J.D., & Armitage, I.M. (1980) Biochemistry (third of three papers in this issue)], permitted the assignment of four histidine residues as ligands to the "catalytic", or A site, metal ions, two coordinated via their N pi imidazole nitrogens and two via N pi. In addition, a fifth histidyl ligand, coordinated through N pi, was shown to be located at the "structural", or B, sites on the dimer. The "regulatory", or C, sites do not contain histidyl metal ligands. Unambiguous identification of the three histidines coordinated to metal ion via N pi was provided by the observation of resolved 113Cd-13C spin-spin coupling (3J = 12-19 Hz) in their gamma-carbon resonances. Once assigned, the 13C resonances of the five histidyl metal ligands were used to monitor the relative affinities of the A, B, and C sites for Cd2+ and Zn2+. At pH 6.3, Cd2+ was found to bind to the A sites at least 10 times tighter than to the B or C sites, which have roughly equal affinities. In marked contrast, Zn2+ was found to have similar affinities for the A and B sites at both pH 6.3 and 8.0. The affinity of the C sites for Zn2+ and Mg2+ was shown to be at least an order of magnitude lower. The binding constants of all three sites for Cd2+ and Zn2+ are greater than 10(5) M-1. Evidence is also presented that suggests that the A, B, and C sites may be located in close proximity to one another in the monomers.
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PMID:Characterization of the properties of the multiple metal binding sites in alkaline phosphatase by carbon-13 nuclear magnetic resonance. 699 14

The structural organization of the epidural lymphatics and lymphatic drainage of the cerebrospinal fluid from spinal meninges was studied in Japanese monkeys (Macaca fuscata) by an enzyme-histochemical method. The spinal meninges were examined at various intervals from 1 to 48 h, as well as at 30 days, following an injection of ultrafine carbon particles into the subarachnoidal space (cisterna magna). Lymphatics were differentiated from blood capillaries by the 5'-nucleotidase (5'-Nase)-alkaline phosphatase (ALPase) double staining method (KATO et al. 1991, 1993) both in the whole-mount preparations and tissue sections. Carbon-filled collecting lymphatics and lymph nodes constantly appeared in the cervical and thoracic regions but only rarely in the lumbo-sacral region after carbon injection. Networks of 5'-Nase-positive lymphatics in the epidural connective tissues were seen in a large area on the dorsal surface around each spinal nerve root in the cervical and upper thoracic regions, especially at a level corresponding to the brachial plexus (C5-Th1). Carbon particles were often found within the 5'-Nase-positive lymphatics. In the lower thoracic and lumbo-sacral regions, on the other hand, the epidural lymphatic network covered only a small area around each spinal nerve root. These findings suggest that the epidural lymphatics are well developed on the dorsal side of the lower cervical spinal dura mater and may function as an absorptive pathway for the cerebrospinal fluid from the subarachnoidal space.
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PMID:Lymphatic drainage of the cerebrospinal fluid from monkey spinal meninges with special reference to the distribution of the epidural lymphatics. 975 4

The present in vitro study investigated select functions (specifically, proliferation, synthesis of intracellular proteins, alkaline phosphatase activity, and deposition of calcium-containing mineral) of osteoblasts (the bone-forming cells) cultured on carbon fibers with nanometer dimensions. Carbon fiber compacts were synthesized to possess either nanophase (i.e., dimensions 100 nm or less) or conventional (i.e., dimensions larger than 100 nm) fiber diameters. Osteoblast proliferation increased with decreasing carbon fiber diameters after 3 and 7 days of culture. Moreover, compared to larger-diameter carbon fibers, osteoblasts synthesized more alkaline phosphatase and deposited more extracellular calcium on nanometer-diameter carbon fibers after 7, 14, and 21 days of culture. The results of the present study provided the first evidence of enhanced long-term (in the order of days to weeks) functions of osteoblasts cultured on nanometer-diameter carbon fibers; in this manner, carbon nanofibers clearly represent a unique and promising class of orthopedic/dental implant formulations with improved osseointegrative properties.
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PMID:Enhanced functions of osteoblasts on nanometer diameter carbon fibers. 1210 99

Carbon nanofibers possess excellent conductivity properties, which may be beneficial in the design of more effective neural prostheses; however, limited evidence on their cytocompatibility properties currently exists. The objective of the present in vitro study was to determine cytocompatibility properties of formulations containing carbon nanofibers pertinent to neural implant applications. Substrates were prepared from four different types of carbon fibers, two with nanoscale diameters (nanophase, or less than or equal to 100 nm) and two with conventional diameters (or greater than 100 nm). Within these two categories, both a high and a low surface energy fiber were investigated and tested. Carbon fibers were compacted in a manual hydraulic press via a uniaxial loading cycle. Astrocytes (glial scar tissue-forming cells) were seeded onto the substrates for adhesion, proliferation, and long-term function studies (such as total intracellular protein and alkaline phosphatase activity). Results provided the first evidence that astrocytes preferentially adhered and proliferated on carbon fibers that had the largest diameter and the lowest surface energy. Based on these results, composite substrates were also formed using different weight percentages (0-25 wt%) of the nanophase, high surface energy fibers in a polycarbonate urethane matrix. Results provided the first evidence of decreased adhesion of astrocytes with increasing weight percents of the high surface energy carbon nanofibers in the polymer composite; this further demonstrates that formulations containing carbon fibers in the nanometer regime may limit astrocyte functions leading to decreased glial scar tissue formation. Positive interactions with neurons, and, at the same time, limited astrocyte functions leading to decreased gliotic scar tissue formation are essential for increased neuronal implant efficacy.
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PMID:Decreased functions of astrocytes on carbon nanofiber materials. 1464 5

A new approach to the modification of carbon nanotubes with biomolecules for the development of nanoscale biosensors is presented. Alkaline phosphatase was immobilized on the surface of multi-wall carbon nanotubes utilizing a layer-by-layer methodology. Carbon nanotubes were incubated with streptavidin, resulting in the formation of a protein layer on the surface of the nanotubes. Biotinylated alkaline phosphatase was then allowed to bind to streptavidin, anchoring the sensing protein onto the surface. Electrochemical biosensors were constructed by using carbon nanotubes compacted into pellets. 1-Naphthyl phosphate, which is hydrolyzed by alkaline phosphatase to the electroactive 1-naphthol, was used as a substrate. Electrodes constructed in this manner were observed to generate an electrochemical signal that was a function of substrate concentration.
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PMID:Protein immobilization on carbon nanotubes through a molecular adapter. 1551 93

Carbon nanotubes (CNTs) have been shown to affect cell behavior. But how and why the CNTs affect potential differentiation of the attached cells has not been largely known. In this study, multiwalled carbon nanotubes (MWNTs) and graphite (GP) were pressed as compacts. Higher ability of CNTs to adsorb proteins, compared with GP, was shown. Myoblastic mouse cells (C2C12) were cultured and the cell responses to the two kinds of compacts were compared in vitro. Meanwhile, we used cell culture on the culture plate as a control. During the conventional culture, significantly better cell attachment, proliferation, and differentiation of cells on the MWNTs were found. To confirm the hypothesis that the larger amount of protein adsorbed on the CNTs was crucial for this, we made the compacts adsorb more proteins in culture medium with 50% fetal bovine serum (FBS) before cell culture. With the adsorption of the proteins in advance, the increments of the total-protein/DNA and alkaline phosphatase (ALP)/DNA for the MWNTs was respectively as about 11 times and 18 times as the increments of those for GP and the control at both day 4 and day 7. Therefore, the CNTs might induce cellular functions by adsorbing more proteins, which indicated that the CNTs might be a candidate for scaffold material for tissue engineering.
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PMID:Effect of carbon nanotubes on cellular functions in vitro. 1876 64

Carbon nanotubes (CNTs) have been synthesized and produced on large scale for their wide application. They have high absorption ability to organic contaminants (such as benzene) and can form CNTs-benzene combination with benzene. In this article, the acute pulmonary toxicity, induced by multiwall carbon nanotubes (MWCNTs), benzene, and their combination, was studied by administrating the three test materials into mice lungs via intratracheal instillation. The biochemical parameters in bronchoalveolar lavage fluid (BALF) and pathological lesions in lungs were used as endpoints to evaluate the pulmonary toxicity of the three test materials at 3-day and 7-day postexposure, respectively. After the mice were intratracheally instilled with MWCNTs, benzene and MWCNTs-benzene combination at doses of 6.67 mg/kg, 2.67 mg/kg, and 9.34 mg/kg (containing 6.67 mg/kg MWCNTs and 2.67 mg/kg benzene), the total protein, alkaline phosphatase (ALP), acid phosphatase (ACP), and lactate dehydrogenase (LDH) in BALF and pathological lesions in lungs were examined. At 3-day postexposure, MWCNTs induced obvious pulmonary toxicity and benzene only induced slight pulmonary toxicity, whereas their combination induced very severe pulmonary toxicity. At 7-day postexposure, MWCNTs and benzene did not induce pulmonary toxicity individually, whereas their combination still induced severe pulmonary toxicity. These data indicated that, at the instilled doses in this experiment, the MWCNTs can alone induce acute pulmonary toxicity in mice and the benzene does not induce pulmonary toxicity, but the pulmonary toxicity of MWCNTs is enhanced after they form MWCNTs-benzene combination with low dose of benzene. The enhanced pulmonary toxicity may be due to the change of MWCNTs aggregation ability after benzene is adsorbed on them.
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PMID:The acute pulmonary toxicity in mice induced by multiwall carbon nanotubes, benzene, and their combination. 1952 38

Carbon nanotubes (CNTs) are a class of new allotrope of carbon. Different functionalized CNTs may vary from their physical and chemical properties to the biological property. In this study, the toxicity of water-soluble taurine multi-walled CNTs (tau-MWNTs), raw MWNTs and positive control crystalline silicon dioxide particles on mouse lungs via intratracheal instillation (i.t.) was investigated. The dosages we used were 0.125, 0.25, 0.5 or 1 mg/kg of tau-MWNTs and raw MWNTs, and 1 mg/kg of silicon dioxide particles; Serum and lungs were collected at 1, 7, 14 or 28 days postexposure. The biochemical and cellular parameters were assessed, which include the ratio of the lung weight and body weight (lung indices), lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and angiotensin converting enzyme (ACE) in serum, and malondialdehyde (MDA), reduced glutathione (GSH), total sulfhydryl group (TSH) in lung tissue homogenates as well as the hydroxyproline in lungs. The characteristic recovery of the lung injury at 28 days postexposure was examined by the assessment of LDH, ALP, lung indices, and histopathology. ACE, MDA, GSH, TSH and histopathological changes showed that tau-MWNTs were less toxic than the raw MWNTs. Histopathological and ultrastructural investigation indicated that the acute pulmonary inflammation in lungs alleviated after 7d postexposure, and were greatly recovered within 28d. Meanwhile, the entrapment of tau-MWNTs reduced greatly by the 28d postexposure. Whereas the heavier pathologic changes induced by raw MWNTs lasted 7 days more than that of tau-MWNTs. Notably, no occurrence of granulomas and fibrosis were found in this study both in the two CNTs samples through 28d postexposure. Silicon dioxide particles, on the contrary, produced more severe damage to lungs than CNTs did in lung index, as well as other biochemical and cellular parameters. These findings indicate that water-soluble tau-MWNTs in low and medium doses induce slight and recoverable pulmonary inflammation in mice, and are less toxic than the insoluble raw MWNTs.
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PMID:Pulmonary toxicity in mice exposed to low and medium doses of water-soluble multi-walled carbon nanotubes. 2112 61

Carbon nanotubes (CNTs), one of the most concerned nanomaterials, with unique electrical, mechanical and surface properties, have been shown suitable for biomedical application. In this study, we evaluated attachment, proliferation, osteogenic gene expression, ALP/DNA, protein/DNA and mineralization of human adipose-derived stem cells cultured in vitro on multi-walled carbon nanotubes (MWNTs) and graphite (GP) compacts with the same dimension. Moreover, we assessed the effect of these two kinds of compacts on ectopic bone formation in vivo. First of all, higher ability of the MWNTs compacts to adsorb proteins, comparing with the GP compacts, was shown. During the conventional culture, it was shown that MWNTs could induce the expression of ALP, cbfa1 and COLIA1 genes while GP could not. Furthermore, alkaline phosphatase (ALP)/DNA and protein/DNA of the cell on the MWNTs compacts, was significantly higher than those of the cells on the GP compacts. With the adsorption of the proteins in culture medium with 50% fetal bovine serum (FBS) in advance, the increments of the ALP/DNA and protein/DNA for the MWNTs compacts were found respectively significantly more than the increments of those for the GP compacts, suggesting that the larger amount of protein adsorbed on the MWNTs was crucial. More results showed that ALP/DNA and protein/DNA of the cells on the two kinds of compacts pre-soaked in culture medium having additional rhBMP-2 were both higher than those of the cells on the samples re-soaked in culture medium with 50% FBS, and that those values for the MWNTs compacts increased much more. Larger mineral content was found on the MWNTs compacts than on the GP compacts at day 7. In vivo experiment showed that the MWNTs could induce ectopic bone formation in the dorsal musculature of ddy mice while GP could not. The results indicated that MWNTs might stimulate inducible cells in soft tissues to form inductive bone by concentrating more proteins, including bone-inducing proteins.
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PMID:The use of carbon nanotubes to induce osteogenic differentiation of human adipose-derived MSCs in vitro and ectopic bone formation in vivo. 2248 42


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