|
pubmed:abstractText |
Vaccinia topoisomerase relaxes DNA through the formation of a covalent DNA-(3'-phosphotyrosyl)protein intermediate at sites containing the sequence 5'-(T/C)CCTT/. The active site, Tyr-274, is situated near the carboxyl terminus of the 314 amino acid enzyme. Here, we report the effects of serial C-terminal deletions. Removal of five amino acids had no effect on topoisomerase activity. However, deletion of 10, 15, or 20 amino acids rendered the enzyme distributive in DNA relaxation, incrementally slowed the rate of single-turnover DNA cleavage, and progressively diminished DNA binding affinity, without altering the sequence specificity of DNA cleavage. These effects lead us to speculate that the region downstream of the active site, which is not well-conserved among the poxvirus-encoded topoisomerases, is a component of the proposed circumferential interface between the enzyme and duplex DNA.
|
