Increased Processivity, Misincorporation, and Nucleotide Incorporation Efficiency in Sulfolobus solfataricus Dpo4 Thumb Domain Mutants.

ABSTRACT: The present study aimed to increase the processivity of Sulfolobus solfataricus DNA polymerase Dpo4. Protein engineering and bioinformatics were used to compile a library of potential Dpo4 mutation sites. Ten potential mutants were identified and constructed. A primer extension assay was used to evaluate the processivity of Dpo4 mutants. Thumb (A181D) and finger (E63K) domain mutants showed a processivity of 20 and 19 nucleotides (nt), respectively. A little finger domain mutant (I248Y) exhibited a processivity of 17 nt, only 1 nt more than wild-type Dpo4. Furthermore, the A181D mutant showed lower fidelity and higher nucleotide incorporation efficiency (4.74 × 10^-4 s^-1 μM^-1) than E63K and I248Y mutants. When tasked with bypassing damage, the A181D m

SUBMITTER: Wang L 

PROVIDER: S-EPMC5583478 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature