Site‐specific insertion of genes into integrons: role of the 59‐base element and determination of the recombination cross‐over point

Abstract

From examination of published DNA sequences of genes found inserted at a specific site in integrons, all genes are shown to be associated, at their 3' ends, with a short imperfect inverted repeat sequence, a 59-base element or relative of this element. The similarity of the arrangement of gene inserts in the integron and in the Tn7 transposon family is described. A refined consensus for the 59-base element is reported. Members of this family are highly diverged and the relationship of a group of longer elements to the 59-base elements is demonstrated. The ability of 59-base elements of different length and sequence to act as sites for recombination catalysed by the integron-encoded DNA integrase is demonstrated, confirming that elements of this family have a common function. The ability of elements located between gene pairs to act as recombination sites has also been demonstrated. The recombination cross-over point has been localized to the GTT triplet which is conserved in the core sites, GTTRRRY, found at the 3' end of 59-base elements. Recombination at the core site found in inverse orientation at the 5' end of the 59-base elements was not detected, and the sequences responsible for orientation of the recombination event appear to reside within the 59-base element. A model for site-specific insertion of genes into integrons and Tn7-like transposons is proposed. Circular units consisting of a gene associated with a 59-base element are inserted into an ancestral element which contains neither a gene nor a 59-base element.(ABSTRACT TRUNCATED AT 250 WORDS)

Gene cassettes: a new class of mobile element

“…The largest group of 59-base elements includes those associated with the aadA genes, aadB, catB3, orfD and aacA (IIa) genes and each of these 59-base elements differs from the consensus for this group (Hall etal., 1991) by only a few bases. Other cases in which closely related 59-base elements are associated with different genes have also been found; for example, the aacA4 and aacA (orfl3) genes encode proteins belonging to distinct protein families but are associated with very similar 59-base elements (see Hall et al, 1991).…”

“…IntIl is believed to be a member of the 1 integrase family on the basis of similarities between the conserved domains shared by members of this family of site-specific recombinases Stokes & Hall, 1989;Abremski & Hoess, 1992). IntIl has been shown to catalyse both integrative and excisive recombination events in in vivo experimental systems (Martinez & de la Cruz, 1990 ;Hall et al, 1991 ;Recchia e t al. , 1994).…”

Comparison of the structure–activity relationships of the integron-associated recombination sites attI3 and attI1 reveals common features

“…These features (Fig. 1a) appear to be generally recognized by IntI-type tyrosine recombinases, as the same 59-be act as sites for several different IntI that are less than 50 % identical at the amino acid sequence level (Martinez & de la Cruz, 1990;Hall et al, 1991;Stokes et al, 1997;Collis et al, 2001Collis et al, , 2002aDrouin et al, 2002; In all attI, potential 7-bp core sites (related to GTTRRRY) at positions corresponding to sites 1 and 2 and separated by 4 or 5 bp, are in bold typescript. Lowercase lettering denotes sequence derived from the first cassette and can vary according to the cassette present.…”

“…1a), suggesting that in this system further molecules of IntI act as accessory factors when bound to the additional sites. The cassette-associated 59-be sites have a set of common features that permit them to be identified, despite substantial sequence diversity and differences in length (Hall et al, 1991;Collis & Hall, 1992a;Stokes et al, 1997). Each 59-be appears to consist of two pairs of inversely oriented IntI-binding sites, each making up a simple site, separated by a segment that has a variable length and sequence but generally includes an inverted repeat (Stokes et al, 1997).…”

“…Gene cassettes are small mobile elements that generally include a single gene and a recombination site known as a 59-be (59-base element) (Hall et al, 1991;Recchia & Hall, 1995. They are normally found in an integron in arrays of one, a few or many cassettes at a specific position which corresponds to the integron-associated attI recombination site (Stokes & Hall, 1989;Hall & Collis, 1995Hall, 2002).…”