A Transcriptional Enhancer Whose Function Imposes a Requirement That Proteins Track Along DNA

Dr, Herendeen; Ga, Kassavetis; Ep, Geiduschek

doi:10.1126/science.1598572

Cited by 130 publications

(111 citation statements)

References 44 publications

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“…These results suggest that the su(Hw) protein does not act as a passive road block for transcription factors; rather, it interacts actively with them either directly, through the leucine zipper region or, indirectly, through other proteins that bind this motif. A requirement for transcription factors to track along the DNA to reach the transcription complex in the promoter has been demonstrated recently for the expression of bacteriophage T4 late genes (Herendeen et al 1992), and this type of mechanism could explain well the directionality in the inhibitory effects of su(Hw).…”

Section: Discussionmentioning

confidence: 99%

DNA position-specific repression of transcription by a Drosophila zinc finger protein.

Geyer¹,

Corcés²

1992

Genes Dev.

406

374

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Section: Discussionmentioning

confidence: 99%

DNA position-specific repression of transcription by a Drosophila zinc finger protein.

Geyer¹,

Corcés²

1992

Genes Dev.

406

374

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“…Various DNA substrates were analyzed, including linear marker DNAs and plasmid DNAs such as pCBR2 and pR67 (4), which contain the gene 16 -17 sequence, and pDH72DE3 (29), a kind gift of Dr. E. P. Geiduschek. Reactions were incubated at 30°C for 3 h and heated at 65°C for 10 min prior to 0.8% agarose gel electrophoresis and ethidium bromide staining following standard methods (25).…”

Section: Methodsmentioning

confidence: 99%

Isolation and Characterization of T4 Bacteriophage gp17 Terminase, a Large Subunit Multimer with Enhanced ATPase Activity

Baumann

Black

2003

Journal of Biological Chemistry

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Phage T4 terminase is a two-subunit enzyme that binds to the prohead portal protein and cuts and packages a headful of concatameric DNA. To characterize the T4 terminase large subunit, gp17 (70 kDa), gene 17 was cloned and expressed as a chitin-binding fusion protein.Following cleavage and release of gp17 from chitin, two additional column steps completed purification. The purification yielded (i) homogeneous soluble gp17 highly active in in vitro DNA packaging (ϳ10% efficiency, >10 8 phage/ml of extract); (ii) gp17 lacking endonuclease and contaminating protease activities; and (iii) a DNA-independent ATPase activity stimulated >100-fold by the terminase small subunit, gp16 (18 kDa), and modestly by portal gp20 and single-stranded binding protein gp32 multimers. Analyses revealed a preparation of highly active and slightly active gp17 forms, and the latter could be removed by immunoprecipitation using antiserum raised against a denatured form of the gp17 protein, leaving a terminase with the increased specific activity (ϳ400 ATPs/gp17 monomer/min) required for DNA packaging. Analysis of gp17 complexes separated from gp16 on glycerol gradients showed that a prolonged enhanced ATPase activity persisted after exposure to gp16, suggesting that constant interaction of the two proteins may not be required during packaging. Many dsDNA1 bacteriophage require the activity of two "terminase" proteins to cut and package concatameric DNA into assembled phage proheads. In T4, the gp16 (small subunit) and gp17 (large subunit) terminase proteins are essential proteins for DNA packaging. (Similar proteins are gpA and gpNu1 in phage, gp18 and gp19 in phage T7/T3, gp2 and gp3 in phage P22, and gp3 and gp16 in phage 29.) As generally found among these phage terminases, the T4 small subunit confers the specific DNA-binding/association properties (1-3). The large subunit contains the prohead-binding and putative DNAtranslocating ATPase activities (3-7). However, because the measured in vitro ATPase activities of isolated terminase proteins (8, 9) have been low in comparison with measured rates of ATP consumption in packaging (10, 11), the identification of terminases as DNA-translocating ATPases has been problematic. What is clear is that packaging is ATP-driven and involves the cooperation of the DNA-associated terminase proteins with the prohead-associated portal vertex protein (7, 12, 13) in phage T4, gp20.The T4 terminase large subunit protein, gp17, has proven difficult to purify because of the protein's limited solubility and toxicity to host cells (14). Coexpression with the gp16 small subunit reduces toxicity; thus, gp17 could be purified with gp16 in low amounts from a temperature-induced expression vector (4). A vector that makes gp17 with a 20-amino acid OmpT leader peptide results in low amounts of uninduced, soluble, and active protein, whereas the overexpressed protein is insoluble and inactive and resists renaturation (15). A His-tagged gp17 protein was also overexpressed and showed similar problems in protein solubi...

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“…In a series of elegant experiments it was demonstrated that gene-45 protein plays an important role in the transcription of genes which are expressed in late stages of infection (Herendeen et al, 1992). The gene-45 protein, in association with the gene-33 and gene-55 proteins, binds to the E. coli RNA polymerase and directs it to the promoters of the phage genes that are expressed in late stages of infection.…”

Section: Rna Transcriptionmentioning

confidence: 99%

PCNA: structure, functions and interactions

1997

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Proliferating cell nuclear antigen (PCNA) plays an essential role in nucleic acid metabolism as a component of the replication and repair machinery. This toroidalshaped protein encircles DNA and can slide bidirectionally along the duplex. One of the wellestablished functions for PCNA is its role as the processivity factor for DNA polymerase d and e. PCNA tethers the polymerase catalytic unit to the DNA template for rapid and processive DNA synthesis. In the last several years it has become apparent that PCNA interacts with proteins involved in cell-cycle progression which are not a part of the DNA polymerase apparatus. Some of these interactions have a direct eect on DNA synthesis while the roles of several other interactions are not fully understood. This review summarizes the structural features of PCNA and describes the diverse functions played by the protein in DNA replication and repair as well as its possible role in chromatin assembly and gene transcription. The PCNA interactions with dierent cellular proteins and the importance of these interactions are also discussed.

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A Transcriptional Enhancer Whose Function Imposes a Requirement That Proteins Track Along DNA

Cited by 130 publications

References 44 publications

DNA position-specific repression of transcription by a Drosophila zinc finger protein.

DNA position-specific repression of transcription by a Drosophila zinc finger protein.

Isolation and Characterization of T4 Bacteriophage gp17 Terminase, a Large Subunit Multimer with Enhanced ATPase Activity

PCNA: structure, functions and interactions

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