E. coli DNA polymerase

T4 DNA polymerase E. coli infected with bacteriophage T4 5 —> 3 chain growth 3 —> 5 exonuclease... 

All DNA synthesis required for DNA repair, recombination, and replication depends on the ability of DNA polymerases to recognize the template and correctly insert the complementary nucleotide. The mechanisms whereby these enzymes achieve this tremendous task has been a central topic of interest since the discovery of the first DNA polymerase, E. coli DNA polymerase I, by Arthur Kornberg approximately half a century ago [1], Since then enormous efforts from scientists in many disciplines have been undertaken to gain insights into the complex mechanisms and functions of these molecular machines. 

The discovery of the first DNA polymerase, E. coli DNA polymerase I, by Kornberg and co-workers occurred over 50 years ago. After extensive research spanning multiple decades, there are currently seven families of polymerases (A, B, C, D, X, Y, and RT) which are classified according to primary sequence homology and structural similarity. Eukaryotic polymerases belong to four of these families (A, B, X, and Y) (reviewed in Shcherbakova et Hubscher et al., Pavlov et al. and McCulloch and Kunkel ). Defects in proper regulation... 

Four incubation mixtures are set up, each containing the DNA template, a specific DNA primer, E. coli DNA polymerase I and all four deoxyribo-nucleoside triphosphates (dNTPs). In addition, each mixture contains a different dideoxynucleoside triphosphate analog, ddATP, ddCTP ddGTP or ddTTP. Incorporation of a dideoxy analog prevents further elongation and so produces a chain termination extension product. The products are electrophoresed on a polyacrylamide gel and the DNA sequence read from the band pattern produced. 

Klenow polymerase (E. coli DNA polymerase I nach Klenow, Boehringer, 2-3 units/ tl)... 

Water soluble carbodiimides inhibit the transcription of supercoiled PM2 DNA with E. coli B RNA polymerase. " Modification of the lactose permease of E. Coli with carbodiimides shows a preference for hydrophobic carbodiimides (DCC) over hydrophilic carbodiimides. In carbodiimide modification of EmrE, a small multidrug transporter in E. Coli, DIPCD modification indicates that Glu-14 is the target of the reaction. Polynucleotides react with positively charged water soluble carbodiimides much faster than do the monomers, owing to their electrostatic effect. ... 

Psoralen interstrand cross-links pose strong blocks to T7 RNA polymerase, E. coli RNA polymerase, and RNA Pol II [54, 65, 81], which is to be expected since the double helix cannot unwind in the vicinity of the adduct. Psoralen monoadducts on the transcribed strand of DNA pose blocks to transcription as well. Historically, psoralen adducts are of great interest since they were among the first chemically induced lesions to be investigated for TC-NER. Interestingly, psoralen interstrand cross-links are subject to TC-NER, but the monoadducts are not [82]. 

Fig, 10.4 Inhibition ofRNA polymerase (E. coli) by proflavine as measured by decreased UMP uptake when DNA, ATP, GDP, and CTP are present in excess. Inset Lineweaver—Burk plot of velocity ofRNA synthesis in proportion to concentration of DNA template supplied (two straight lines one with and one without proflavine) Hurwitz et ai, 1962). 

Superscript II cDNA synthesis kit containing RT (200U/ jL), 5x first-strand reaction buffer, O.IM DTT, lOmM dNTP mix, Superscript II RT, 5x second-strand reaction buffer, E. coli DNA ligase, E. coli DNA polymerase I, E. coli RNase H, T4 DNA polymerase, and DEPC-treated water. (Invitrogen 11917-010). 

Correct transcription of phage DNA by E. coli RNA polymerase needs holoenzyme (Burgess et al., 1969 Hinkle and Chamberlin, 1970). In... 

Transcription of natural templates like T4 and TZ DNA by E, coli RNA polymerase yields defined RNA products at both high and low ionic strength and without the help of additional factors (Millette et al., 1970). At the ionic strength of the in vitro protein synthesizing systems (preincubated S30 or... 

DNA-dependent RNA polymerase E. coli) Core enzyme 5 0.732 72P1... 

B Guenther, R Onrust, A Sail, M O Donnell, J Kuriyan. Crystal structure of the 5 subunit of the clamp-loader complex of E. coli DNA polymerase III. Cell 91 335-345, 1997. 

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