Complete transposition requires four active monomers in the mu transposase tetramer.
A tetramer of Mu transposase (MuA) cleaves and joins multiple DNA strands to promote transposition. Derivatives of MuA altered at two acidic residues that are conserved among transposases and retroviral integrases form tetramers but are defective in both cleavage and joining. These mutant proteins were used to analyze the contribution of individual monomers to the activity of the tetramer. The performance of different protein combinations demonstrates that not all monomers need to be catalytically competent for the complex to promote an individual cleavage or joining reaction. Furthermore, the results indicate that each pair of essential residues is probably donated to the active complex by a single monomer. Although stable, tetramers composed of a mixture of mutant and wild-type MuA generate products cleaved at only one end and with only one end joined to the target DNA. The abundance of these abortive products and the ratios of the two proteins in complexes stalled at different steps indicate that the complete reaction requires the activity of all four monomers. Thus, each subunit of MuA appears to use the conserved acidic amino acids to promote one DNA cleavage or one DNA joining reaction.
Department of Biology, Massachusetts Institute of Technology, Cambridge.
T A
TA
Baker
E
E
Kremenstova
L
L
Luo
D010583:Bacteriophage mu / Q000201:enzymology* / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D004254:DNA Nucleotidyltransferases / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism; D005814:Genes, Viral; D019426:Integrases; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation; D009713:Nucleotidyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D045522:Recombinases; D019895:Transposases
D004277:DNA, Single-Stranded; D004279:DNA, Viral; D045522:Recombinases; D004254:DNA Nucleotidyltransferases; D019426:Integrases; D009713:Nucleotidyltransferases; D019895:Transposases; C115006:integron integrase IntI1