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)

The parameters governing the activity of the cloned T4 gene 23, which codes for the major T4 head protein, were analyzed. Suppressor-negative bacteria carrying wild-type T4 gene 23 cloned into plasmid pCR1 or pBR322 were infected with T4 gene 23 amber phage also carrying mutations in the following genes: (i) denA and denB (to prevent breakdown of plasmid DNA after infection) and (ii) denA, denB, and, in addition, 56 (to generate newly replicated DNA containing dCMP) and alc/unf (because mutations in this last gene allow late genes to be expressed in cytosine-containing T4 DNA). Bacteria infected with these phage were labeled with (14)C-amino acids at various times after infection, and the labeled proteins were separated by one-dimensional gel electrophoresis so that the synthesis of plasmid-coded gp23 could be compared with the synthesis of other, chromosome-coded T4 late proteins. We analyzed the effects of additional mutations that inactivate DNA replication proteins (genes 32 and 43), an RNA polymerase-binding protein (gene 55), type II topoisomerase (gene 52), and an exonuclease function involved in recombination (gene 46) on the synthesis of plasmid-coded gp23 in relation to chromosome-coded T4 late proteins. In the denA:denB:56:alc/unf genetic background, the phage chromosome-borne late genes followed the same regulatory rules (with respect to DNA replication and gp55 action) as in the denA:denB genetic background. The plasmid-carried gene 23 was also under gp55 control, but was less sensitive than the chromosomal late genes to perturbations of DNA replication. Synthesis of plasmid-coded gp23 was greatly inhibited when both the type II T4 topoisomerase and the host's DNA gyrase are inactivated. Synthesis of gp23 was also substantially affected by a mutation in gene 46, but less strongly than in the denA:denB genetic background. These observations are interpreted as follows. The plasmid-borne T4 gene 23 is primarily expressed from a late promoter. Expression of gene 23 from this late promoter responds to an activation event which involves some structural alteration of DNA. In these respects, the requirements for expressing the plasmid-borne gene 23 and chromosomal late genes are very similar (although in the denA:denB:56:alc/unf genetic background, there are significant quantitative differences). For the plasmid-borne gene 23, activation involves the T4 gp46, a protein which is required for DNA recombination. However, for the reasons presented in the accompanying paper (Jacobs et al., J. Virol. 39:31-45, 1981), we conclude that the activation of gene 23 does not require a complete breakage-reunion event which transposes that gene to a later promoter on the phage chromosome. Ways in which gp46 may actually be involved in late promoter activation on the plasmid are discussed.
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PMID:Regulation of expression of cloned bacteriophage T4 late gene 23. 626 20

A preparation of bacteriophage T4-induced deoxyribonucleotide synthetase complex is described. This very large complex of enzymes can be separated by centrifugation at 100,000 X g, by sucrose step gradient centrifugation, or with molecular exclusion columns. By direct assay and by unidimensional and two-dimensional acrylamide electrophoretic separations the following T4-coded enzymes were shown to be associated with the complex: ribonucleoside diphosphate reductase, dCMP deaminase, dCTP/dUTPase, dCMP hydroxymethylase, dTMP synthetase, and DNA polymerase. Other phage-coded prereplicative proteins related to DNA replication and other phage functions such as the proteins coded by genes 32, 46, rIIA, and rIIB as well as many unidentified proteins were also consistently associated with the isolated fractions. T4 DNA topoisomerase, a membrane-bound enzyme, was found in quantity in all purified fractions of the complex, even in preparations apparently free of membrane and of T4 DNA. The functional integrity of a segment of the complex was followed by measuring the conversion of [5-3H]CDP to the level of 5-hydroxymethyl dCMP. This series of reactions requires the actions of T4-coded ribonucleoside diphosphate reductase and its associated reducing system, dCTP/dUTPase and dCMP hydroxymethylase, 3H being lost to water at the last step. In this reaction sequence an intermediate, [5-3H]dCMP, is maintained at low steady state concentrations, and argument is presented that the synthesis of deoxyribonucleotides is channeled and normally tightly coupled to DNA replication. One of the primary characteristics of this complex is its ready dissociation of dilution into smaller complexes of proteins and to the free forms of the proteins. That the complex is held together by weak electrostatic forces was supported by its sensitivity to dissociation at moderate salt concentrations. Not only the enzymes required in deoxyribonucleotide synthesis but T4 DNA polymerase, T4 DNA topoisomerase, and a number of other proteins dissociate to varying degrees from the larger complexes under these conditions.
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PMID:Characteristics of a bacteriophage T4-induced complex synthesizing deoxyribonucleotides. 675 52