title: Receipt of the C-terminal tail from a neighboring lambda Int protomer allosterically stimulates Holliday junction resolution.
abstract: Bacteriophage lambda integrase (Int) catalyzes the integration and excision of the phage lambda chromosome into and out of the Esherichia coli host chromosome. The seven carboxy-terminal residues (C-terminal tail) of Int comprise a context-sensitive regulatory element that links catalytic function with protein multimerization and also coordinates Int functions within the multimeric recombinogenic complex. The experiments reported here show that the beta5-strand of Int is not simply a placeholder for the C-terminal tail but rather exerts its own allosteric effects on Int function in response to the incoming tail. Using a mutant integrase in which the C-terminal tail has been deleted (W350ter), we demonstrate that the C-terminal tail is required for efficient and accurate resolution of Holliday junctions by tetrameric Int. Addition of a free heptameric peptide of the same sequence as the C-terminal tail partially reverses the W350ter defects by stimulating Holliday junction resolution. The peptide also stimulates the topoisomerase function of monomeric W350ter. Single residue alterations in the peptide sequence and a mutant of the beta5 strand indicate that the observed stimulation arises from specific contacts with the beta5 strand (residues 239-243). The peptide does not stimulate binding of W350ter to its cognate DNA sites and therefore appears to recapitulate the effects of the normal C-terminal tail intermolecular contacts in wild-type Int. Models for the allosteric stimulation of Int activity by beta5 strand contacts are discussed.
authors.affiliation: Department of Molecular Biology, Cell Biology, and Biochemistry, Brown University, Providence, RI 02912, USA.
authors.forename: Dane
authors.initials: D
authors.lastname: Hazelbaker
authors.forename: Marta
authors.initials: M
authors.lastname: Radman-Livaja
authors.forename: Arthur
authors.initials: A
authors.lastname: Landy
mesh_terms: D000495:Allosteric Site; D010582:Bacteriophage lambda / Q000201:enzymology* / Q000235:genetics; D002876:Chromosomes, Bacterial / Q000378:metabolism; D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry; D045566:DNA, Cruciform; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D019426:Integrases / Q000737:chemistry* / Q000235:genetics*; D008957:Models, Genetic; D008958:Models, Molecular; D008968:Molecular Conformation; D009690:Nucleic Acid Conformation; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry; D011995:Recombination, Genetic; D013997:Time Factors
chemical_list: D045566:DNA, Cruciform; D010455:Peptides; D011506:Proteins; D004247:DNA; D019426:Integrases