Cleavage of the Site-Specific Recombination Protein γ δ Resolvase: The Smaller of Two Fragments Binds DNA Specifically

• Published in: Proceedings of the National Academy of Sciences - PNAS 1984-04, Vol.81 (7), p.2001-2005
• Main Authors: Abdel-Meguid, Sherin S., Nigel D. F. Grindley, Templeton, Nancy Smyth, Steitz, Thomas A.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amino acids; Base Sequence; Binding Sites; Biochemistry; Chymotrypsin; Crossovers; Crystals; DNA; Electrophoresis; Gels; Genetic equilibrium; Molecular Weight; Monomers; Nucleic Acid Conformation; Nucleotidyltransferases - metabolism; Peptide Fragments - analysis; Protein Binding; resolvase; Transposases; X-ray crystallography
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.81.7.2001

The 20,500-dalton γ δ resolvase monomer can be cleaved by chymotrypsin into a 5000-dalton COOH-terminal fragment and a 15,500-dalton NH2-terminal fragment that have been purified. Two crystal forms of the large fragment have been obtained, one of which is isomorphous with crystals of the native protein, showing that the large fragment makes the protein--protein contacts in the crystal and that the small fragment is segmentally disordered relative to the large fragment. Nuclease protection demonstrates that the small fragment binds specifically to all three DNA binding sites protected by resolvase. However, unlike native resolvase, which binds to all three complete sites with equal affinity, the small fragment binds to each of the six half sites with a different affinity. It has not been possible to demonstrate specific DNA binding of the larger fragment. Thus, resolvase has a modular construction analogous to that found for some repressors and activators; its COOH-terminal domain recognizes specific sequences in the DNA and its NH2-terminal domain mediates protein--protein interactions and probably has the enzymatic activity.