Purification of a DNA topoisomerase II from the hyperthermophilic archaeon Sulfolobus shibatae. A thermostable enzyme with both bacterial and eucaryal features

• Published in: The Journal of biological chemistry Vol. 269; no. 44; pp. 27663 - 27669
• Main Authors: Bergerat, A, Gadelle, D, Forterre, P
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 04.11.1994; American Society for Biochemistry and Molecular Biology
• Subjects: DNA Topoisomerases, Type II - chemistry; DNA Topoisomerases, Type II - isolation & purification; DNA Topoisomerases, Type II - metabolism; Hot Temperature; Molecular Weight; Protein Conformation; Protein Denaturation; Substrate Specificity; Sulfolobus - enzymology; Sulfolobus shibatae; Topoisomerase II Inhibitors
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)47037-8

A type II DNA topoisomerase has been purified to homogeneity from the hyperthermophilic archaeon Sulfolobus shibatae. The enzyme is composed of two subunits of 60 and 47 kDa. It has a Stokes radius of 69 A and has a sedimentation coefficient of 7.8 S which gives a calculated native molecular mass of approximately 230 kDa, indicating a heterotetrameric structure. This enzyme is ATP and Mg2+ dependent and can relax both negatively and positively supercoiled DNA, but presents no supercoiling activity. The S. shibatae DNA topoisomerase II is more efficient in decatenation than in relaxation. The optimal temperature for the enzymatic activity is approximately 80 degrees C. This archaeal enzyme is not inhibited by the gyrase inhibitor novobiocin but is sensitive to several inhibitors of eucaryotic DNA topoisomerases of type II such as amsacrines, ellipticine, and the quinolone CP-115,953. Like all prokaryotic DNA topoisomerase II, the S. shibatae DNA topoisomerase II is a heterotetramer but the absence of supercoiling activity, the strong decatenase activity, and the pattern of antibiotic sensitivity of the S. shibatae DNA topoisomerase II is reminiscent of eucaryotic enzymes.