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:5.99.1.2 (topoisomerase)
9,166 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

DNA topoisomerase II is an essential nuclear enzyme required for the proper condensation and segregation of chromosomes during mitotic and meiotic cell division. The enzyme exists in the cell as a phosphoprotein, and it is most highly phosphorylated in G2 and M-phases of the cell cycle. We have shown that topoisomerase II is the target of casein kinase II (CKII) in yeast by comparison of in vivo and in vitro phosphotryptic peptide maps. Limited proteolysis and probing with domain specific antibodies show that with the exception of a weakly modified residue between aa 660 and aa 1250, all residues modified by CKII are in the last 200 amino acids of yeast topoisomerase II. This C-terminal domain is the least conserved region of the enzyme and truncation of the enzyme shows that it is nonessential for activity in vitro. However, the fully dephosphorylated full-size protein is nearly inactive in decatenation assays, and activity can be restored by phosphorylation by CKII. To reconcile these observations, we propose that the C-terminal region is a negative regulatory domain, counteracted by phosphorylation within the domain itself. To test this hypothesis we have mutagenised 12 potential CKII phosphoacceptor sites in the C-terminus of topoisomerase II and introduced the mutant genes into a yeast strain which has a temperature sensitive top2 gene. The growth of the transformed strains is monitored at nonpermissive temperature to determine whether C-terminal phosphorylation is important for mitotic growth. In addition, we have purified the mutant enzymes to homogeneity for in vitro assays.
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PMID:The regulation of DNA topoisomerase II by casein kinase II. 773 31

Eukaryotic DNA topoisomerase II is an abundant nuclear enzyme that is essential for cell proliferation. This homodimeric enzyme catalyzes the cleavage and re-ligation of double-stranded DNA required to separate replicated sister chromatids. Both biochemical and genetic studies show that its catalytic activity is required for chromosome condensation and segregation, and that its decatenation activity can be stimulated by a variety of protein kinases in vitro. In budding yeast, topoisomerase II is most highly phosphorylated in metaphase, and casein kinase II (CKII) was shown to be the major kinase modifying topoisomerase II. We have investigated the effects of phosphorylation of yeast topoisomerase II by CKII in vitro, by means of gel-retardation and filter binding assays. The phosphorylation of the C terminus of topoisomerase II by CKII appears to increase the stability of the complex formed with linear DNA fragments, while dephosphorylation has the opposite effect. Rephosphorylation of phosphatase-treated topoisomerase II by chicken casein kinase II restores a stable protein-DNA complex using a linear DNA fragment. The enhanced stability of the topoisomerase II-DNA complex is also observed with relaxed circular DNA, but not with supercoiled minicircles, in agreement with published results using topoisomerase II from Drosophila. Limited proteolysis and probing with domain-specific antibodies shows that, with the exception of a weakly modified residue between amino acid residues 660 and 1250, all residues modified by casein kinase II are in the last 180 amino acid residues of yeast topoisomerase II.
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PMID:Phosphorylation of the C-terminal domain of yeast topoisomerase II by casein kinase II affects DNA-protein interaction. 793 31

We present a novel assay for the study of protein-protein interactions involving DNA topoisomerase II. Under various conditions of incubation we observe that topoisomerase II forms complexes at least tetrameric in size, which can be sedimented by centrifugation through glycerol. The multimers are enzymatically active and can be visualized by electron microscopy. Dephosphorylation of topoisomerase II inhibits its multimerization, which can be restored at least partially by rephosphorylation of multiple sites within its 200 C-terminal amino acids by casein kinase II. Truncation of topoisomerase II just upstream of the major phosphoacceptor sites reduces its aggregation, rendering the truncated enzyme insensitive to either kinase treatments or phosphatase treatments. This is consistent with a model in which interactions involving the phosphorylated C-terminal domain of topoisomerase II aid either in chromosome segregation or in chromosome condensation.
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PMID:Topoisomerase II forms multimers in vitro: effects of metals, beta-glycerophosphate, and phosphorylation of its C-terminal domain. 793 13

Topoisomerase II protein is essential for cell proliferation and is known to exist as a phosphoprotein in cells from both lower and higher eukaryotic species. In this paper, we have investigated the phosphorylation of the alpha isozyme of human topoisomerase II. The topoisomerase II alpha protein was phosphorylated predominantly on serine residues in the human tumor cell lines HeLa and NSCLC-3. Two-dimensional tryptic phosphopeptide mapping studies revealed several sites of phosphorylation in vivo, including a major site that was common to topoisomerase II alpha protein from both HeLa and NSCLC-3 cells. To identify sites of phosphorylation, the regulatory C-terminal domain of human topoisomerase II alpha protein was overexpressed in Escherichia coli as a hexahistidine-tagged fusion protein and purified by nickel chelate chromatography. Tryptic phosphopeptide mapping revealed that casein kinase II phosphorylated the C-terminal domain primarily on 2 serine residues in vitro, which were shown to be sites of modification in vivo. Site-directed mutagenesis studies identified these casein kinase II-specific phosphorylation sites as serine 1524 and serine 1376.
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PMID:Serine 1524 is a major site of phosphorylation on human topoisomerase II alpha protein in vivo and is a substrate for casein kinase II in vitro. 796 67

Two sublines of LY murine lymphoma, differing in sensitivity to CPT, served as source of topoisomerase I in order to compare the enzyme's properties. The activity of topoisomerase I isolated from LY-S cells of reduced sensitivity to CPT increased about 2-times more upon phosphorylation with casein kinase but was inhibited to a lesser extent upon dephosphorylation with alkaline phosphatase than the enzyme from the CPT-sensitive LY-R cells. The in vitro phosphorylation of LY-S enzyme restored its sensitivity to CPT. The in vitro incorporation of 32P into topoisomerase protein was about 1.7-times higher in LY-S than in LY-R enzyme. A reversed incorporation ratio was observed upon metabolic labelling. The level of topoisomerase I protein, determined by Western blot analysis using scleroderma anti-topoisomerase I antibodies, was about 1.5-times higher in LY-S than in LY-R cells. The level of topoisomerase I mRNA was similar in both sublines. These results indicate that the reduced sensitivity of LY-S cells to CPT is based on the lowered phosphorylation of topoisomerase I protein but does not depend on the expression of topoisomerase I gene.
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PMID:Topoisomerase I is differently phosphorylated in two sublines of L5178Y mouse lymphoma cells. 799 92

The nucleotide sequence of a 55098 bp region from the right end of the genome of a virulent African swine fever virus (ASFV) isolate (Malawi LIL20/1) has been determined. Translation of the sequence identified 67 major open reading frames (ORFs) which are closely spaced and read from both DNA strands. At six positions intergenic tandem repeat arrays are found. Comparison of the predicted amino acid sequences of encoded proteins with protein sequence databases identified a number of homologies. These include three subunits of RNA polymerase, a protein with homology to transcription factor SII (TFSII), a DNA ligase, two subunits of mRNA capping enzyme, a DNA topoisomerase type II, a dUTPase, a protein kinase, three helicases, a ubiquitin-conjugating enzyme, a protein with homology to the nif S and nif S-like proteins identified in some bacteria and Saccharomyces cerevisiae, a protein with homology to both a myeloid differentiation primary response antigen (MyD116) and to a herpes simplex virus-encoded neurovirulence-associated protein (ICP34.5), a protein with homology to the ASFV-encoded structural protein p22, two proteins with homology to copies of the ASFV-encoded multigene family 360 and one protein with homology to the ASFV-encoded multigene family 110. Four genes encode proteins which have homology to each other and constitute a new multigene family (MGF100). Nine ORFs encode proteins which contain predicted transmembrane domains. The possible functions of these predicted ASFV-encoded proteins are discussed and the evolutionary relationship of ASFV to other viruses are considered. Despite the similarities in genome structure and replication strategy of ASFV with poxviruses, sequence similarity between them is low and the organization of ASFV-encoded genes is not colinear with that of the orthopoxviruses.
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PMID:Nucleotide sequence of a 55 kbp region from the right end of the genome of a pathogenic African swine fever virus isolate (Malawi LIL20/1). 802 96

The activity of several proteins involved in the development of antitumor drug resistance is regulated by protein phosphorylation. These proteins include the mdr-1-encoded P-glycoprotein (Pgp) and topoisomerase II (topo II). The corresponding evidence is reviewed and attempts to modulate multidrug resistance (MDR) by protein kinase C inhibitors are described. The expression of several proteins which are essential in drug resistance is regulated at the transcriptional level, involving protein phosphorylation by members of the protein kinase C (PKC) family, casein kinase II (CKII), and others. These proteins include mdr-1-encoded P-glycoprotein, metallothionein, glutathione S-transferase (GST), dTMP synthase, and the proteins Fos and Jun. The corresponding genes are under positive regulation of ras, which in turn requires the activation of a protein kinase cascade for its function. Protein kinases are therefore potentially useful targets in reducing the expression of proteins involved in the development of multifactorial drug resistance caused by the expression of transforming ras-genes. Attempts to inhibit the ras-induced fos expression by an inhibitor of protein kinase C (ilmofosine) are described. Protein kinase inhibitors are also able to synergistically enhance the cytotoxicity of cis-platinum, which is discussed as resulting from a reduction of PKC-dependent fos expression.
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PMID:Role of protein kinases in antitumor drug resistance. 806 Nov 7

Bufalin, an active principle of the traditional Chinese medicine chan'su, has been proved to be a potent differentiation inducer in human leukemia cells. To study the mechanism of the differentiation of human leukemia ML1 cells induced by bufalin, we measured the effect of 10 nM bufalin on cell growth, activities of various protein kinases, and cell cycle. The ML1 cell growth was inhibited significantly at 24 hr and the inhibiting effect persisted for 6 days. Activities of PKC, PKA, cdc2 kinase and CK II in ML1 cells were changed early by bufalin; PKA and PKC activities were inhibited, and cdc2 kinase and CK II activities were increased. These results suggest that bufalin induces differentiation of ML1 cells by modulating several protein kinase activities in a distinct way from RA and 1 alpha, 25(OH) 2D3. Cell cycle changes, measured by flow cytometry, became evident at 12 hr after treatment of ML1 cells with bufalin and the cells were preferentially arrested in the G2/M phase. This effect of bufalin on the cell cycle of leukemia cells is similar to that of topoisomerase inhibitors. Indeed, the activity of topoisomerase II but not topoisomerase I of ML1 cells was inhibited remarkably by the treatment of the cells with 10 nM bufalin.
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PMID:Cell cycle arrest and protein kinase modulating effect of bufalin on human leukemia ML1 cells. 807 71

Immunoprecipitation of DNA topoisomerase II from yeast results in a preparation that contains casein kinase II; this suggests that the two proteins may associate in the intact cell. Purified recombinant topoisomerase II and casein kinase II associate to form a complex in vitro which is stable after topoisomerase II becomes phosphorylated by the kinase. Studies with isolated recombinant casein kinase II subunits disclosed that although the alpha (catalytic) subunit alone can efficiently phosphorylate topoisomerase II, the formation of a stable topoisomerase II-casein kinase II association requires the presence of the beta subunit of the kinase. Both proteins engaged in this complex retain their catalytic activities. Naturally occurring polyamines and polyanionic compounds appear to be crucial factors governing the interaction between the two proteins. Although the biological significance of a stable catalytically active topoisomerase II-casein kinase II molecular complex remains to be defined, these observations suggest the possibility of a novel mechanism regulating topoisomerase II and casein kinase II activities.
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PMID:DNA topoisomerase II and casein kinase II associate in a molecular complex that is catalytically active. 822 2

Topoisomerase I (Topo I) is involved in many cellular functions that involve unwinding of supercoiled DNA, such as transcription and replication. Topo I is also the target of autoimmune antibodies in progressive systemic sclerosis (scleroderma), and abnormal regulation of Topo I may influence the excessive production of collagen found in scleroderma. Topo I is phosphorylated in vivo at serine residues and, in vitro, the activity of Topo I is increased by phosphorylation by casein kinase type II (CKII) and protein kinase C (PKC). In this study, a protein kinase activity from rat liver nuclei is shown to copurify with Topo I during Bio-Rex 70 cation exchange chromatography. The kinase can phosphorylate Topo I at serine residues, resulting in a threefold increase in topoisomerase activity. A relatively tight association between this kinase and Topo I is demonstrated by the ability to coprecipitate the kinase with scleroderma autoimmune anti-Topo I antibodies. The kinase activity is similar to CKII since it is Ca2+ and cyclic nucleotide independent, it can utilize either ATP or GTP as phosphate donor, and it can phosphorylate casein and phosvitin, but not histones. However, unlike typical CKII, the Topo I-associated kinase could utilize Mn2+ almost as well as Mg2+, it is not stimulated by polyamines, and it does not appear to undergo autophosphorylation. In conclusion, we demonstrate that rat liver Topo I is relatively tightly associated with a CKII-like protein kinase that can phosphorylate and activate Topo I. These findings provide corroborative evidence that CKII, or a CKII-like protein kinase, is a physiologic regulator of Topo I.
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PMID:A casein kinase type II (CKII)-like nuclear protein kinase associates with, phosphorylates, and activates topoisomerase I. 826 Jan 98


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