Probing the binding of coumarins and cyclothialidines to DNA gyrase

DNA gyrase is the target of a number of antibacterial agents, including the coumarins and the cyclothialidines. To extend our understanding of the mechanism of action of these compounds, we have examined the previously published crystal structures of the complexes between the 24 kDa fragment of GyrB and coumarin and cyclothialidine drugs and made mutations by site-directed mutagenesis. We used proteolysis as a probe of drug binding to wild-type and mutant proteins. Limited proteolysis of gyrase revealed that binding of these antibiotics is associated with a characteristic proteolytic fingerprint, suggesting a drug-induced conformational change. The ability of the mutants to bind the drugs was studied by testing their ability to induce the coumarin-associated proteolytic signature and to bind to a novobiocin-affinity column. To analyze further the interaction of the drugs with gyrase, we studied the binding using surface plasmon resonance. Mutation of Asn46 to Asp has only a modest effect on the binding of coumarins, while an Asn46 to Leu mutation results in a 10-fold decrease in the affinity. Mutation of Asp73 to Asn completely abolishes binding to both coumarins and cyclothialidines. Mutations at these residues also abolish ATP hydrolysis, explaining the inability of such mutations to occur spontaneously.     

Sex steroids,ANGELS and osteoporosis

Osteoporosis is characterized by reduced bone density and strength. Bone mass peaks between age 30 and 40 and then declines. This can be accelerated by factors including menopause and insufficient dietary calcium. Hormone replacement therapy (HRT) is currently the standard treatment for osteoporosis. However, growing concern over potential side effects of HRT has driven a search for alternative therapies. A recent report 1 reveals a potential alternative to HRT: a gender-neutral synthetic steroid that increases bone mass and strength without affecting reproductive organs. This compound acts via a novel extranuclear sex steroid receptor signaling mechanism that has important implications for nuclear receptor biology and human health.     

Establishment Of Comperiella Bifasciata (Hym.: Encyrtidae) On Aonidiella Aurantii (Hom.: Diaspididae) In Cyprus

Reference is made to the introduction, rearing, colonization and establishment of Comperiella bifasciata Howard to improve the effectiveness of biological control of the California red scale,Aonidiella aurantii (Maskell), in Cyprus. The parasitoid was colonized during 1985- 89 and in 1993- 94 it was recovered in all the surveyed areas. It has been permanently established and is more abundant in orchards with microclimatic conditions characteristic of intermediate and interior areas whereAphytis melinus DeBach is the dominant red scale parasite.Aphytis chrysomphali (Mercet) prevails only in some of the coastal areas with mild and rather humid microclimates. In the surveyed orchards the overall adult relative abundance of the three main parasitoids, A. melinus, A. chrysomphali and C. bifasciata was 54.1%, 42.1%, and 3.8%, respectively.     

Evidence for a conformational change in the DNA gyrase-DNA complex from hydroxyl radical footprinting.

We have used the technique of hydroxyl radical footprinting to probe the complex between DNA gyrase and a 198 bp DNA fragment containing the preferred gyrase cleavage site from plasmid pBR322. We find that gyrase protects 128 bp from the hydroxyl radical with the central 13 bp (adjacent to the gyrase cleavage site) being most strongly protected. Flanking the central region are arms showing periodic protection from the reagent suggesting a helical repeat of 10.6 bp, consistent with the DNA being wrapped upon the enzyme surface. The presence of 5'-adenylyl-beta,gamma-imidodiphosphate or a quinolone drug causes alteration of the protection pattern consistent with a conformational change in the complex involving one arm of the wrapped DNA. The significance of these results for the mechanism of DNA supercoiling by gyrase is discussed.     

A unified theory of gene expression

The human genome has been called "the blueprint for life." This master plan is realized through the process of gene expression. Recent progress has revealed that many of the steps in the pathway from gene sequence to active protein are connected, suggesting a unified theory of gene expression.     

Control ofSaissetia oleae (Hom.: Coccidae) in Cyprus through establishment ofMetaphycus barletti andM. Helvolus (Hym.: Encyrtidae)

In Cyprus the black scale,Saissetia oleae (Olivier) is a pest primarily of olive,Olea europaea L., but it also attacks several other plants, among which areCitrus spp. and oleander,Nerium oleander L. The parasitoidsMetaphycus bartletti Annecke &; Mynhardt andMetaphycus helvolus (Compere) have been imported from France, massreared and permanently established in Cyprus. Following limited releases of these parasitoids, populations ofS. oleae have been reduced from outbreak levels to almost non-existence and remained at such low levels even after discontinuation of these releases.