title: Constraints for zinc finger linker design as inferred from X-ray crystal structure of tandem Zif268-DNA complexes.
abstract: Zinc-finger proteins offer a versatile and effective framework for the recognition of DNA binding sites. By connecting multiple fingers together with canonical TGEKP linkers, a protein may be designed to recognize almost any desired target DNA sequence. However, proteins containing more than three zinc-fingers do not bind as tightly as one might predict, and it appears that some type of strain is introduced when a six-finger protein is constructed with canonical linkers. In an attempt to understand the sources of this strain, we have solved the 2.2A resolution X-ray crystallographic structure of a complex that has two copies of the three-finger Zif268 protein bound to adjacent sites on one duplex DNA. Conceptually, this is equivalent to a six-finger protein in which the central linker has been removed and the complex has been allowed to "relax" to its most stable conformation. As in other Zif268-DNA complexes, the DNA is approximately linear and is slightly underwound. Surprisingly, the structure of the complex is similar (within 0.5A) to an arrangement that would allow a canonical linker at the center of the complex, and it seems possible that entropic effects (involving the librational degrees of freedom in the complex) could be important in determining optimal linker length.
authors.affiliation: Department of Biology, Howard Hughes Medical Institute, Cambridge, MA, USA. epeisach@med-xtal.bu.edu
authors.forename: Ezra
authors.initials: E
authors.lastname: Peisach
authors.forename: Carl O
authors.initials: CO
authors.lastname: Pabo
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D046911:Macromolecular Substances; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016335:Zinc Fingers
chemical_list: D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D009692:Nucleic Acid Heteroduplexes; D011994:Recombinant Proteins; D014157:Transcription Factors; D004247:DNA