Identification of the region that determines the specificity of binding of the transposases encoded by Tn3 and gamma delta to the terminal inverted repeat sequences.
To analyze the region that determines the specificity of binding of the Tn3 transposase to the terminal inverted repeat sequences (IR), we first determined the nucleotide sequence of a Tn3-family transposon, gamma delta, which is supposed to encode a transposase similar to that of Tn3. gamma delta was 5981 bp in length and contained three coding frames: Two were the genes, tnpA and tnpR, encoding transposase (1002 amino acids) and resolvase/repressor (183 amino acids), respectively, and the third, named tnpX, encoding a protein (698 amino acids) of unknown function but containing two NTP-binding motifs. Utilizing the tnpA sequence, we then constructed a series of Tn3-gamma delta hybrid genes encoding chimeric proteins in the N-terminal segments of the transposases (amino acid position 1 to 242 of Tn3 or 1' to 243' of gamma delta), which has been previously shown to be responsible for specific binding of transposase to IR sequences in Tn3. Examination of their DNA-binding activities revealed that the subsegment of the N-terminus from amino acid position 1 to 109 determines the specificity of binding to the IR sequences. The third coding frame found in gamma delta, tnpX, is located downstream of tnpR and is expressed from the tnpR promoter in the absence of the tnpR gene product, resolvase/repressor, to produce a protein that inhibits the growth of the host cells. Possible roles of this protein are discussed.
Maekawa
T
T
Institute of Molecular and Cellular Biosciences, University of Tokyo, Japan.
Ohtsubo
E
E
D000595:Amino Acid Sequence; D000668:Ampicillin Resistance / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D004251:DNA Transposable Elements / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005374:Filtration; D005798:Genes, Bacterial / Q000235:genetics; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics*; D012097:Repressor Proteins / Q000235:genetics; D017422:Sequence Analysis, DNA; D019895:Transposases
D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D009713:Nucleotidyltransferases; D019895:Transposases