期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2003
卷号：100
期号：13
页码：7509-7514
DOI：10.1073/pnas.0832468100
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Members of the transposase/retroviral-integrase superfamily use a single active site to perform at least two reactions during transposition of a DNA transposon or a retroviral cDNA. They hydrolyze a DNA sequence at the end of the mobile DNA and then join this DNA end to a target DNA (a reaction called DNA strand transfer). Critical to understanding the mechanism of recombination is elucidating how these distinct reactions are orchestrated by the same active site. Here we find that DNA substrates terminating in a dideoxynucleotide allow Mu transposase to hydrolyze a target DNA, combining aspects of both natural reactions. Analyses of the sequence preferences for target hydrolysis and of the structure of the cleaved product indicate that this reaction is promoted by the active site in the conformation that normally promotes DNA strand transfer. Dissecting the DNA requirements for target hydrolysis reveals that the ribose of the last nucleotide of the Mu DNA activates transposase's catalytic potential, even when this residue is not a direct chemical participant. These findings provide insight into the molecular mechanism insuring that DNA strand transfer ordinarily occurs rather than inappropriate DNA cleavage. The required presence of the terminal nucleotide in the transposase active site creates a great advantage for the attached 3'OH to serve as nucleophile.
