Consequences of producing DNA gyrase from a syntheticgyrBAoperon inSalmonella entericaserovar Typhimurium

Abstract

AbstractDNA gyrase is an essential type II topoisomerase that is composed of two subunits, GyrA and GyrB and has an A2B2structure. Although both subunits are required in equal proportions to form DNA gyrase, thegyrAandgyrBgenes that encode them inSalmonella(and in many other bacteria) are at widely separated locations on the chromosome, are under separate transcriptional control and are present in different copy numbers in rapidly growing bacteria (gyrAis near the terminus of chromosome replication whilegyrBis near the origin). We generated a syntheticgyrBAoperon at theoriC-proximal location ofgyrBto test the significance of the gyrase gene position forSalmonellaphysiology. Producing gyrase from an operon did not alter growth kinetics, cell morphology, competitive fitness index, or sensitivity to some gyrase-inhibiting antibiotics. However, the operon strain had altered DNA supercoiling set points, its SPI-2 virulence genes were expressed at a reduced level and its survival was reduced in macrophage. ThegyrBgene could not be deleted from itsoriC-proximal location, even in agyrBmerodiploid strain. We discuss the physiological significance of the differentgyrAandgyrBgene arrangements found naturally inSalmonellaand other bacteria.