Identification of a topoisomerase IV in actinobacteria: purification and characterization of ParYR and GyrBR from the coumermycin A1 producer Streptomyces rishiriensis DSM 40489.
The biosynthetic gene clusters of the gyrase inhibitors coumermycin A(1) and clorobiocin contain two different resistance genes (gyrB(R) and parY(R)). Both genes code for B subunits of type II topoisomerases. The authors have now overexpressed and purified the encoded proteins, as well as the corresponding A subunits GyrA and ParX. Expression was carried out in Streptomyces lividans in the form of hexahistidine fusion proteins, allowing purification by nickel affinity chromatography. The complex of GyrA and GyrB(R) was found to catalyse ATP-dependent supercoiling of DNA, i.e. to function as a gyrase, whereas the complex of ParX and ParY(R) catalysed ATP-dependent decatenation and relaxation, i.e. the functions of topoisomerase IV (topo IV). This is believed to represent the first topo IV identified in the class of actinobacteria, and the first demonstration of the formation of a topo IV as a resistance mechanism of an antibiotic producer.
Eberhard-Karls-Universität Tübingen, Pharmazeutische Biologie, Auf der Morgenstelle 8, D-72076 Tübingen, Germany.
Elisabeth
E
Schmutz
Eberhard-Karls-Universität Tübingen, Pharmazeutische Biologie, Auf der Morgenstelle 8, D-72076 Tübingen, Germany.
Susanne
S
Hennig
Eberhard-Karls-Universität Tübingen, Pharmazeutische Biologie, Auf der Morgenstelle 8, D-72076 Tübingen, Germany.
Shu-Ming
SM
Li
Eberhard-Karls-Universität Tübingen, Pharmazeutische Biologie, Auf der Morgenstelle 8, D-72076 Tübingen, Germany.
Lutz
L
Heide
D039903:Actinobacteria / Q000201:enzymology* / Q000235:genetics; D049933:Aminocoumarins; D000900:Anti-Bacterial Agents / Q000096:biosynthesis; D001483:Base Sequence; D003374:Coumarins / Q000378:metabolism* / Q000494:pharmacology; D027081:DNA Gyrase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D027101:DNA Topoisomerase IV / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D005810:Multigene Family; D009154:Mutation; D009690:Nucleic Acid Conformation; D021122:Protein Subunits; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics; D013696:Temperature
D049933:Aminocoumarins; D000900:Anti-Bacterial Agents; D003374:Coumarins; D004269:DNA, Bacterial; D021122:Protein Subunits; D027101:DNA Topoisomerase IV; D027081:DNA Gyrase; C004628:coumermycin