title: Binding linkage in a telomere DNA-protein complex at the ends of Oxytricha nova chromosomes.
abstract: Alpha and beta protein subunits of the telomere end binding protein from Oxytricha nova (OnTEBP) combine with telomere single strand DNA to form a protective cap at the ends of chromosomes. We tested how protein-protein interactions seen in the co-crystal structure relate to DNA binding through use of fusion proteins engineered as different combinations of domains and subunits derived from OnTEBP. Joining alpha and beta resulted in a protein that bound single strand telomere DNA with high affinity (K(D-DNA)=1.4 nM). Another fusion protein, constructed without the C-terminal protein-protein interaction domain of alpha, bound DNA with 200-fold diminished affinity (K(D-DNA)=290 nM) even though the DNA-binding domains of alpha and beta were joined through a peptide linker. Adding back the alpha C-terminal domain as a separate protein restored high-affinity DNA binding. The binding behaviors of these fusion proteins and the native protein subunits are consistent with cooperative linkage between protein-association and DNA-binding equilibria. Linking DNA-protein stability to protein-protein contacts at a remote site may provide a trigger point for DNA-protein disassembly during telomere replication when the single strand telomere DNA must exchange between a very stable OnTEBP complex and telomerase.
authors.affiliation: Department of Biology, University of Utah, 257 S 1400 E, Salt Lake City, UT 84112-0840, USA.
authors.forename: Pawel
authors.initials: P
authors.lastname: Buczek
authors.forename: Rochelle S
authors.initials: RS
authors.lastname: Orr
authors.forename: Sean R
authors.initials: SR
authors.lastname: Pyper
authors.forename: Mili
authors.initials: M
authors.lastname: Shum
authors.forename: Emily
authors.initials: E
authors.lastname: Kimmel
authors.forename: Irene
authors.initials: I
authors.lastname: Ota
authors.forename: Shawn E
authors.initials: SE
authors.lastname: Gerum
authors.forename: Martin P
authors.initials: MP
authors.lastname: Horvath
mesh_terms: D000818:Animals; D002875:Chromosomes / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D046912:Multiprotein Complexes; D016813:Oxytricha / Q000737:chemistry / Q000235:genetics*; D011485:Protein Binding; D021122:Protein Subunits; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D016615:Telomere / Q000378:metabolism*; D034501:Telomere-Binding Proteins / Q000378:metabolism*
chemical_list: D046912:Multiprotein Complexes; D021122:Protein Subunits; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D034501:Telomere-Binding Proteins; D004247:DNA