Purification and characterization of a novel ATP-independent type I DNA topoisomerase from a marine methylotroph.
DNA topoisomerase is involved in DNA repair and replication. In this study, a novel ATP-independent 30-kDa type I DNA topoisomerase was purified and characterized from a marine methylotroph, Methylophaga sp. strain 3. The purified enzyme composed of a single polypeptide was active over a broad range of temperature and pH. The enzyme was able to relax only negatively supercoiled DNA. Mg(2+) was required for its relaxation activity, while ATP gave no effect. The enzyme was clearly inhibited by camptothecin, ethidium bromide, and single-stranded DNA, but not by nalidixic acid and etoposide. Interestingly, the purified enzyme showed Mn(2+)-activated endonuclease activity on supercoiled DNA. The N-terminal sequence of the purified enzyme showed no homology with those of other type I enzymes. These results suggest that the purified enzyme is an ATP-independent type I DNA topoisomerase that has, for the first time, been characterized from a marine methylotroph.
Kwack
Man Sup
MS
Department of Biotechnology and Research Center for Proteineous Materials, Chosun University, Gwangju 501-759, Republic of Korea.
Park
Jung Eun
JE
Park
Jong Kun
JK
Lee
Jung Sup
JS
D000255:Adenosine Triphosphate / Q000494:pharmacology; D002413:Cations, Divalent / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D004264:DNA Topoisomerases, Type I / Q000145:classification / Q000302:isolation & purification* / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D004996:Ethidium / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008345:Manganese / Q000494:pharmacology; D008386:Marine Biology; D044147:Piscirickettsiaceae / Q000145:classification* / Q000201:enzymology*; D013997:Time Factors; D059004:Topoisomerase I Inhibitors
D002413:Cations, Divalent; D004277:DNA, Single-Stranded; D059004:Topoisomerase I Inhibitors; D008345:Manganese; D000255:Adenosine Triphosphate; D004492:Edetic Acid; D004264:DNA Topoisomerases, Type I; D004996:Ethidium