Characteristics of DNA polymerase I from an extreme thermophile, Thermus scotoductus strain K1

The DNA polymerase from the extreme thermophileThermus scotoductus strain K1 (TsK1) was expressed inEscherichia coli, purified, and characterized. The enzyme demonstrated an optimal temperature and pH value of 72 –74°C and 9.0, respectively, and could efficiently amplify 2.5 kb DNA products.TsK1 DNA polymerase did not require additional K+ ions but it did need Mg2+ at 3 –5 mM for optimal activity. The base insertion fidelity for this enzyme was significantly better than that ofTaq DNA polymerase. AbstractSeveral native and engineered heat ‐stable DNA polymerases from a variety of sources are used as powerful tools in different molecular techniques, including polymerase chain reaction, medical diagnostics, DNA sequencing, biological diversity assessments, and in vitro mutagenesis. The DNA polymerase from the extreme thermophile,Thermus scotoductus strain K1, (TsK1) was expressed inEscherichia coli, purified, and characterized. This enzyme belongs to a distinct phylogenetic clade, different from the commonly used DNA polymerase I enzymes, including those fromThermus aquaticus andThermus thermophilus. The enzyme demonstrated an optimal temperature and pH value of 72 –74°C and 9.0, respectively, and could efficiently amplify 2.5 kb DNA products.TsK1 DNA polymerase did not require additional K+ ions but it did need Mg2+ at 3 –5 mM for optimal activity. It was stable for at least 1 h at 80°C, and its half‐life at 88 and 95°C was 30 and 15 min, respectively. ...
Source: MicrobiologyOpen - Category: Microbiology Authors: Tags: ORIGINAL ARTICLE Source Type: research