Defining the structural and functional roles of the carboxyl region of the bacteriophage lambda excisionase (Xis) protein.
The bacteriophage lambda excisionase (Xis) protein is required for excisive site-specific recombination. Xis is composed of 72 amino acids and binds cooperatively to two DNA sites (X1 and X2) that are arranged as direct repeats. Alternatively, Xis binds cooperatively with the host-encoded factor for inversion stimulation (FIS) protein at the X1 and F sites, respectively. Here we analyzed the effects of missense substitutions from codon 57 to the carboxyl end of the protein and nonsense mutations that truncate the protein at various positions from residues 60 to 69. We find that all of the mutant proteins promote excision to some extent and interact cooperatively with FIS. Some mutants have no detectible phenotype while others are altered in their abilities to promote excision or to interact cooperatively with integrase (Int). Computer modeling predicts that amino acids from residues 59 to 65 are in an alpha-helix conformation. Mutants with substitutions on one side of the helix at residues 57, 60, 63 and 64 as well as truncated mutants containing 60, 61 or 63 amino acids, fail to interact cooperatively with Int suggesting that this region of the protein forms the interface with Int. Mutants with substitutions at other positions in the putative helix have no detectible phenotype. Residues 66 to 68 may form a reverse turn and the last four amino acids (69 to 72) may not be crucial for the structure or function of the protein.
Wu
Z
Z
Department of Microbiology, College of Medicine, University of Illinois, Urbana, IL 61801, USA.
Gumport
R I
RI
Gardner
J F
JF
D000595:Amino Acid Sequence; D010582:Bacteriophage lambda / Q000201:enzymology*; D001483:Base Sequence; D003001:Cloning, Molecular; D003198:Computer Simulation; D004254:DNA Nucleotidyltransferases / Q000737:chemistry* / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry; D008242:Lysogeny / Q000502:physiology; D008969:Molecular Sequence Data; D016296:Mutagenesis / Q000235:genetics; D009876:Operon / Q000235:genetics; D010446:Peptide Fragments / Q000235:genetics; D010641:Phenotype; D017433:Protein Structure, Secondary; D011995:Recombination, Genetic / Q000235:genetics; D014764:Viral Proteins / Q000737:chemistry
D004268:DNA-Binding Proteins; D010446:Peptide Fragments; D014764:Viral Proteins; D004254:DNA Nucleotidyltransferases; C063911:excisionase