Control of Strand Scission by Type IIA Topoisomerases

摘要: Topoisomerases are a family of essential enzymes that disentangle chromosomes and manage DNA supercoils, targets broad class successful antibiotics anticancer therapeutics. The type IIA topoisomerases operate by complex ATP-dependent strand passage mechanism in which the enzyme transports one segment through transient, enzyme-mediated break second segment. To understand cleavage topoisomerases, I used suicide substrate to crystallize binding core covalently bound its active-site catalytic tyrosine. crystal structure revealed II IA employ novel variation canonical two-metal ion phosphoryl-transfer chemistry achieve cleavage. Additionally, enabled me determine first fully-catalytic topo II-DNA-nucleotide complex. overall doubly-domain swapped architecture enzyme. This organization produces an unexpected interaction between conformational transducing element ATPase domain, is critical for both DNA-stimulated ATP hydrolysis global topoisomerase activity. data indicate domains pivot about each other ensure unidirectional this state senses promote turnover reset. Lastly, DNA-bound two human isoforms, IIα, underscores coupling active site configuration with metal occupancy at center, as well dimerization status key dissociable interface over 50 A away. also highlights amino acid differences drug-binding pocket isoforms could serve differentiating features developing more selective anti-topoisomerase agents. work presented dissertation helps explain years biochemical studies, unifies many elements control allostery, has implications understanding large DNA-remodeling molecular machines whole, means small molecules target these clinical benefit.