Isolation of temperature-sensitive DNA polymerase III from Saccharomyces cerevisiae cdc2-2

Biochemistry ◽  
1991 ◽  
Vol 30 (32) ◽  
pp. 8092-8096 ◽  
Author(s):  
Ann Blank ◽  
Lawrence A. Loeb
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Polymerase ◽  
Temperature Sensitive ◽  
Dna Polymerase Iii ◽  
Polymerase Iii

The Mechanism of Replication of Single-Stranded DNA. VI. Requirements for Ribonucleoside Triphosphates and DNA Polymerase III

1973 ◽  
Vol 51 (12) ◽  
pp. 1588-1597 ◽  
Author(s):  
David T. Denhardt ◽  
Makoto Iwaya ◽  
Grant McFadden ◽  
Gerald Schochetman
Keyword(s):  
Dna Polymerase ◽  
Dna Polymerases ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Dna Polymerase Iii ◽  
Single Stranded Dna ◽  
E Coli ◽  
Polymerase Iii

Evidence is presented that in Escherichia coli made permeable to nucleotides by exposure to toluene, the synthesis of a DNA chain complementary to the infecting single-stranded DNA of bacteriophage [Formula: see text] requires ATP as well as the four deoxyribonucleoside triphosphates. This synthesis results in the formation of the parental double-stranded replicative-form (RF) molecule. The ATP is not required simply to prevent degradation of the ribonucleoside or deoxyribonucleoside triphosphates; it can be partially substituted for by other ribonucleoside triphosphates.No single one of the known E. coli DNA polymerases appears to be uniquely responsible in vivo for the formation of the parental RF. Since [Formula: see text] replicates well in strains lacking all, or almost all, of the in-vitro activities of DNA polymerases I and II, neither of these two enzymes would seem essential; and in a temperature-sensitive E. coli mutant (dnaEts) deficient in DNA polmerase-I activity and possessing a temperature-sensitive DNA polymerase III, the viral single-stranded DNA is efficiently incorporated into an RF molecule at the restrictive temperature. In contrast, both RF replication and progeny single-stranded DNA synthesis are dependent upon DNA polymerase III activity.

scholarly journals INTERACTION OF DNA POLYMERASE III γ AND β SUBUNITS IN VIVO IN SALMONELLA TYPHIMURIUM

Genetics ◽  
1986 ◽  
Vol 113 (3) ◽  
pp. 499-515
Author(s):  
Joyce Engstrom ◽  
Annette Wong ◽  
Russell Maurer
Keyword(s):  
Salmonella Typhimurium ◽  
Dna Polymerase ◽  
Regulatory Gene ◽  
Temperature Sensitive ◽  
Physical Interaction ◽  
Dna Polymerase Iii ◽  
Β Subunit ◽  
Γ Subunit ◽  
Polymerase Iii

ABSTRACT We show that temperature-sensitive mutations in dnaZ, the gene for the γ subunit of DNA polymerase III holoenzyme, can be suppressed by mutations in the dnaN gene, which encodes the β subunit. These results support a direct physical interaction of these two subunits during polymerase assembly or function. The suppressor phenotype is also sensitive to modulation by the dnaA genotype. Since dnaA is organized in an operon with dnaN, and dnaA is a regulatory gene of this operon, we propose that the dnaA effect on suppression can best be explained by modulation of suppressor dnaN levels.

scholarly journals The second subunit of DNA polymerase III ( ) is encoded by the HYS2 gene in Saccharomyces cerevisiae

1998 ◽  
Vol 26 (2) ◽  
pp. 477-485 ◽  
Author(s):  
K. Hashimoto ◽  
N. Nakashima ◽  
T. Ohara ◽  
S. Maki ◽  
A. Sugino
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Polymerase ◽  
Dna Polymerase Iii ◽  
Polymerase Iii

scholarly journals Suppression of Temperature-Sensitive Chromosome Replication of an Escherichia coli dnaX(Ts) Mutant by Reduction of Initiation Efficiency

2003 ◽  
Vol 185 (12) ◽  
pp. 3583-3595 ◽  
Author(s):  
Alexandra Blinkova ◽  
Mary Jo Hermandson ◽  
James R. Walker
Keyword(s):  
Dna Polymerase ◽  
Temperature Sensitivity ◽  
Temperature Sensitive ◽  
Dna Polymerase Iii ◽  
Suppressor Mutations ◽  
Polymerase Iii ◽  
Initiation Frequency ◽  
Cold Sensitive ◽  
Ts Mutant

ABSTRACT Temperature sensitivity of DNA polymerization and growth of a dnaX(Ts) mutant is suppressible at 39 to 40°C by mutations in the initiator gene, dnaA. These suppressor mutations concomitantly cause initiation inhibition at 20°C and have been designated Cs,Sx to indicate both phenotypic characteristics of cold-sensitive initiation and suppression of dnaX(Ts). One dnaA(Cs,Sx) mutant, A213D, has reduced affinity for ATP, and two mutants, R432L and T435K, have eliminated detectable DnaA box binding in vitro. Two models have explained dnaA(Cs,Sx) suppression of dnaX, which codes for both the τ and γ subunits of DNA polymerase III. The initiation deficiency model assumes that reducing initiation efficiency allows survival of the dnaX(Ts) mutant at the somewhat intermediate temperature of 39 to 40°C by reducing chromosome content per cell, thus allowing partially active DNA polymerase III to complete replication of enough chromosomes for the organism to survive. The stabilization model is based on the idea that DnaA interacts, directly or indirectly, with polymerization factors during replication. We present five lines of evidence consistent with the initiation deficiency model. First, a dnaA(Cs,Sx) mutation reduced initiation frequency and chromosome content (measured by flow cytometry) and origin/terminus ratios (measured by real-time PCR) in both wild-type and dnaX(Ts) strains growing at 39 and 34°C. These effects were shown to result specifically from the Cs,Sx mutations, because the dnaX(Ts) mutant is not defective in initiation. Second, reduction of the number of origins and chromosome content per cell was common to all three known suppressor mutations. Third, growing the dnaA(Cs,Sx) dnaX(Ts) strain on glycerol-containing medium reduced its chromosome content to one per cell and eliminated suppression at 39°C, as would be expected if the combination of poor carbon source, the Cs,Sx mutation, the Ts mutation, and the 39°C incubation reduced replication to the point that growth (and, therefore, suppression) was not possible. However, suppression was possible on glycerol medium at 38°C. Fourth, the dnaX(Ts) mutation can be suppressed also by introduction of oriC mutations, which reduced initiation efficiency and chromosome number per cell, and the degree of suppression was proportional to the level of initiation defect. Fifth, introducing a dnaA(Cos) allele, which causes overinitiation, into the dnaX(Ts) mutant exacerbated its temperature sensitivity.

DNA polymerase III is required for DNA repair in Saccharomyces cerevisiae

1993 ◽  
Vol 24 (3) ◽  
pp. 200-204 ◽  
Author(s):  
Waldemar Suszek ◽  
Hanna Baranowska ◽  
Jerzy Zuk ◽  
Witold J. Jachymczyk
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Repair ◽  
Dna Polymerase ◽  
Dna Polymerase Iii ◽  
Polymerase Iii

scholarly journals Replication-Dependent Recruitment of the β-Subunit of DNA Polymerase III from Cytosolic Spaces to Replication Forks in Escherichia coli

2002 ◽  
Vol 184 (3) ◽  
pp. 867-870 ◽  
Author(s):  
Toshinari Onogi ◽  
Katsufumi Ohsumi ◽  
Tsutomu Katayama ◽  
Sota Hiraga
Keyword(s):  
Escherichia Coli ◽  
Dna Polymerase ◽  
Chromosome Replication ◽  
Temperature Sensitive ◽  
Replication Forks ◽  
Dna Polymerase Iii ◽  
Β Subunit ◽  
Nonpermissive Temperature ◽  
Polymerase Iii

ABSTRACT The β-subunit of DNA polymerase III is located as one or two condensed clusters within the nucleoid-occupied space in exponentially growing cells of Escherichia coli. When chromosome replication is terminated after incubation at nonpermissive temperature in a temperature-sensitive dnaC mutant, the β-subunit is located in the cytosolic spaces of the cell poles.

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