Escherichia coli thioredoxin confers processivity on the DNA polymerase activity of the gene 5 protein of bacteriophage T7.

Abstract:

Bacteriophage T7 gene 5 protein has been purified to apparent homogeneity from cells overexpressing its gene several hundred-fold. Gene 5 protein is a DNA polymerase with low processivity; it dissociates from the primer-template after catalyzing the incorporation of 1-50 nucleotides, depending on the salt concentration. Escherichia coli thioredoxin, a host protein that is tightly associated with the gene 5 protein in phage-infected cells, is not required for this activity. Thioredoxin acts as an accessory protein to bestow processivity on the polymerizing reaction; DNA synthesis catalyzed by the gene 5 protein-thioredoxin complex on a single-stranded DNA template can polymerize thousands of nucleotides without dissociation. Conditions that increase the stability of secondary structures in the template (i.e., low temperature or high ionic strength) decrease the processivity. E. coli single-stranded DNA-binding protein stimulates both the rate of elongation and the processivity of the gene 5 protein-thioredoxin complex.

Polymerases:

Topics:

Historical Protein Properties (MW, pI, ...), Kinetic Parameters, Nucleotide Incorporation, Source / Purification

Status:

new topics/pols set partial results complete validated

Results:

Polymerase Reference Property Result Context
T7 (no thioreoxin) Tabor S1987 Molecular Weight 8E+04 Dalton Technique: SDS-PAGE
T7 (no thioreoxin) Tabor S1987 Cloned or native Cloned in E. coli
T7 (no thioreoxin) Tabor S1987 Full length or truncated Full length
T7 (no thioreoxin) Tabor S1987 Processivity 1bp
T7 (no thioreoxin) Tabor S1987 Specific Activity 110 units/mg Technique: Polymerase Assay (calf thymus DNA)
T7 (no thioreoxin) Tabor S1987 Specific Activity 14 units/mg Technique: Polymerase Assay (M13 DNA)
T7 Tabor S1987 Cloned or native Cloned in E. coli
T7 Tabor S1987 Tagged No
T7 Tabor S1987 Full length or truncated Full length
T7 Tabor S1987 Processivity 10kb
T7 Tabor S1987 Specific Activity 8300 units/mg Technique: Polymerase Assay (calf thymus DNA)
T7 Tabor S1987 Specific Activity 5.3E+04 units/mg Technique: Polymerase Assay (M13 DNA)
T7 Tabor S1987 Vmax 100 /second Reaction: Nucleotide incorporation; Substrate: dNTPs
Eco Pol I Tabor S1987 Full length or truncated Full length
Eco Pol I Tabor S1987 Specific Activity 9800 units/mg Technique: Polymerase Assay (calf thymus DNA)
Eco Pol I Tabor S1987 Specific Activity 2.8E+04 units/mg Technique: Polymerase Assay (M13 DNA)

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