Reprogramming the purine nucleotide cofactor requirement of Drosophila P element transposase in vivo.
Guanosine triphosphate (GTP)-binding proteins are involved in controlling a wide range of fundamental cellular processes. In vitro studies have indicated a role for GTP during Drosophila P element transposition. Here we show that P element transposase contains a non-canonical GTP-binding domain that is critical for its ability to mediate transposition in Drosophila cells. Moreover, a single amino acid substitution could switch the nucleotide binding-specificity of transposase from GTP to xanthosine triphosphate (XTP). Importantly, this mutant protein could no longer function effectively in transposition in vivo but required addition of exogenous xanthine or xanthosine for reactivation. These results suggest that transposition may be controlled by physiological GTP levels and demonstrate that a single mutation can switch the nucleotide specificity for a complex cellular process in vivo.
Mul
Y M
YM
Department of Molecular and Cell Biology, University of California, Berkeley 94720-3204, USA.
Rio
D C
DC
D000818:Animals; D002460:Cell Line; D016384:Consensus Sequence; D004254:DNA Nucleotidyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004251:DNA Transposable Elements; D004330:Drosophila / Q000201:enzymology* / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D006160:Guanosine Triphosphate / Q000378:metabolism*; D015151:Immunoblotting; D009154:Mutation; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding; D012263:Ribonucleosides / Q000494:pharmacology; D012265:Ribonucleotides / Q000378:metabolism*; D019895:Transposases; D019820:Xanthine; D014970:Xanthines / Q000494:pharmacology
D004251:DNA Transposable Elements; D012263:Ribonucleosides; D012265:Ribonucleotides; D014970:Xanthines; D019820:Xanthine; C065326:xanthosine 5'-triphosphate; D006160:Guanosine Triphosphate; C005893:xanthosine; D004254:DNA Nucleotidyltransferases; D019895:Transposases