Trans-acting transposase mutant from Tn5.
Transposition of Tn5 and of its component insertion sequence IS50R is regulated through the action of two proteins it encodes: a cis-acting transposase, Tnp, and a trans-acting inhibitor of transposition, Inh. The mechanism of the cis-acting Tnp and the relevance of inhibition to cis action have been addressed in the current study. A specific colony morphology assay for transposition of Tn5 was shown to be sensitive to Inh produced in trans and was used to screen for mutants in Inh and/or Tnp with altered regulation. A dominant mutant in IS50R that promotes transposition in trans was isolated and characterized. The mutant (449F) carries a Leu----Phe mutation at position 449 in Tnp. This mutation reduces the frequency of Tn5 or IS50R transposition in cis but allows Tnp-449F to act as efficiently in trans as it does in cis. Tnp-449F is sensitive to inhibition and, furthermore, Inh-449F is a competent inhibitor in trans. These results show that Tnp-449F is a trans-acting transposase, unlike wild-type Tnp, which is cis-acting.
DeLong
A
A
Department of Biology, Yale University, New Haven, CT 06511.
Syvanen
M
M
D010582:Bacteriophage lambda / Q000235:genetics; D004251:DNA Transposable Elements; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D005838:Genotype; D009154:Mutation; D009713:Nucleotidyltransferases / Q000235:genetics*; D010641:Phenotype; D010957:Plasmids; D015183:Restriction Mapping; D019895:Transposases
D004251:DNA Transposable Elements; D009713:Nucleotidyltransferases; D019895:Transposases