Transposition of mobile DNA elements generates genetic diversity. Advantage has been taken of this process in the development of transposable elements as tools for insertional mutagenesis (Granger et al.
), transgenesis (Robinson et al.
) and as gene-therapy vectors (Ivics & Izsvak, 2006
; Wilson et al.
The eukaryotic mariner
is a member of the Tc1/mariner
family of transposable elements and was first isolated from Drosophila mauritiana
elements are found in the genomes of most eukaryotes, including the human genome (Robertson & Zumpano, 1997
moves by a cut-and-paste mechanism involving a series of DNA-hydrolysis and transesterification reactions related to the integration of retroviruses (such as HIV-1) and the RAG1/2 recombinase-mediated rearrangement of immunoglobulin genes.
The sole requirement for Mos1
transposition in vitro
is the Mos1 transposase, a 40 665 Da enzyme encoded by a single gene within the 1.3 kbp element. In the first step of transposition, the transposase recognizes the 28-base-pair DNA inverted repeats (IRs) at either end of the element. This interaction is sequence-specific and is mediated by the N-terminal DNA-binding domain of the transposase (Zhang et al.
; Augé-Gouillou et al.
). Subsequently, the transposon ends are brought together to form a paired-ends complex (PEC) and the transposon is excised from flanking DNA. It has previously been shown (Dawson & Finnegan, 2003
) that the 5′-end of the nontransferred strand (NTS) is nicked three bases within the element, whereas the 3′-end of the transferred strand (TS) is cleaved precisely at the junction of the transposon and flanking DNA. In the final steps, target DNA is recruited to the PEC and the transposon DNA is inserted at the 5′-end of a TA dinucleotide.
The structure of the bacterial Tn5 transposase with transposon DNA (Davies et al.
) provided the first picture of a transposition intermediate. However, less is known of the structures of eukaryotic transposases or the complexes that they form with DNA. The structure of the bipartite DNA-binding domain of Tc3 transposase was determined in complex with IR DNA (Watkins et al.
) and more recently the structure of the apo form of the N-terminally truncated eukaryotic hAT transposase Hermes was solved (Hickman et al.
). Previously, we determined the structure of the catalytic domain of Mos1 transposase (Richardson et al.
) and showed that two divalent metal ions could bind in the active site. Invoking a two-metal mechanism for DNA hydrolysis, we argued that PEC intermediates containing protein dimers were sufficient to catalyse all the steps of Mos1
Here, we report the crystallization of a complex formed between full-length Mos1 transposase and its inverted-repeat DNA. The crystals diffract X-rays to 3.25 Å and the data display twofold noncrystallographic symmetry (NCS). These results are discussed in relation to proposed models of the PEC.