title: The additional 165 amino acids in the B protein of Escherichia coli DNA gyrase have an important role in DNA binding.
abstract: DNA gyrase is the only enzyme known to negatively supercoil DNA. The enzyme is a heterotetramer of A(2)B(2) subunit composition. Alignment of the primary sequence of gyrase B (GyrB) from various species shows that they can be grouped into two classes. The GyrB of Gram-negative eubacteria has a stretch of about 165 amino acids in the C-terminal half, which is lacking in other GyrB subunits and type II topoisomerases. In Escherichia coli, no function has so far been attributed to this stretch. In this study, we have tried to assess the function of this region both in vivo and in vitro. A deletant (GyrBDelta160) lacking this region is non-functional in vivo. The holoenzyme reconstituted from gyrase A (GyrA) and GyrBDelta160 shows reduced but detectable supercoiling and quinolone-induced cleavage activity in vitro. GyrBDelta160 retains its ability to bind to GyrA and novobiocin. However, when reconstituted with GyrA, the deletant shows greatly impaired DNA binding. The intrinsic ATPase activity of the GyrBDelta160 is comparable to that of wild type GyrB, but this activity is not stimulated by DNA. These studies indicate that the additional stretch present in GyrB is essential for the DNA binding ability of E. coli gyrase.
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors.forename: M
authors.initials: M
authors.lastname: Chatterji
authors.forename: S
authors.initials: S
authors.lastname: Unniraman
authors.forename: A
authors.initials: A
authors.lastname: Maxwell
authors.forename: V
authors.initials: V
authors.lastname: Nagaraja
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D001483:Base Sequence; D017931:DNA Primers; D004250:DNA Topoisomerases, Type II / Q000737:chemistry / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology*; D011485:Protein Binding; D017384:Sequence Deletion
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial; D000251:Adenosine Triphosphatases; D004250:DNA Topoisomerases, Type II