Crystallization and structure determination of the core-binding domain of bacteriophage lambda integrase.
Bacteriophage lambda integrase catalyzes site-specific DNA recombination. A helical bundle domain in the enzyme, called the core-binding domain (Int(CB)), promotes the catalysis of an intermediate DNA-cleavage reaction that is critical for recombination and is not well folded in solution in the absence of DNA. To gain structural insights into the mechanism behind the accessory role of this domain in catalysis, an attempt was made to crystallize an Int(CB)-DNA complex, but crystals of free Int(CB) were fortuitously obtained. The three-dimensional structure of DNA-free Int(CB) was solved at 2.0 A resolution by molecular replacement using as the search model the previously available DNA-bound 2.8 A structure of the Int(CB) domain in a larger construct of lambda integrase. The crystal structure of DNA-free Int(CB) resembles the DNA-bound structure of Int(CB), but exhibits subtle differences in the DNA-binding face and lacks electron density for ten residues in the C-terminus that form a portion of a linker connecting Int(CB) to the C-terminal catalytic domain of the enzyme. Thus, this work reveals the domain in the absence of DNA and allows comparison with the DNA-bound form of this catalytically activating domain.
Biophysics Program and Department of Biochemistry, The Ohio State University, Columbus, OH 43201, USA.
Hari B
HB
Kamadurai
Rinku
R
Jain
Mark P
MP
Foster
D000595:Amino Acid Sequence; D010582:Bacteriophage lambda / Q000201:enzymology*; D002384:Catalysis; D020134:Catalytic Domain; D003460:Crystallization; D018360:Crystallography, X-Ray; D004269:DNA, Bacterial / Q000378:metabolism; D019426:Integrases / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009994:Osmolar Concentration; D011485:Protein Binding; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D014961:X-Ray Diffraction
D004269:DNA, Bacterial; D019426:Integrases