Inter-replicon transposition of Tn1/3 occurs in two sequential genetically separable steps

The 4,957-base pair (bp) transposon Tn3 encodes three polypeptides, two of which are needed for normal inter-replicon transposition and one, β-lactamase, which is responsible for the transposon's ampicillin resistance (Ap^r)^1,2. The closely related transposon Tn1 specifies three interchangeable polypeptides of identical function. The 1,015-amino acid tnpA protein is essential for transposition, whereas the 185-residue tnpR protein regulates the rate of transposition^3,4 and is required for the generation of normal end products of inter-replicon transposition^5,6. Here we report the inter-replicon transposition properties of tnpA^- tnpR^- derivatives of both Tn1 and Tn3. We show that the tnpA product is sufficient to generate a transpositional co-integrate intermediate in which the donor and recipient replicons become fused between two directly repeated copies of the transposon. In the absence or presence of tnpA product, such co-integrates are efficiently converted to normal transposition end products in the presence of tnpR protein. Two amber mutants selected because of their derepressed tnpR-mediated control of transposition are also unable to resolve co-integrate intermediates in a Sup⁰ (suppressor minus) background. For each mutant, both mutant phenotypes are suppressed by SupD and SupE but not by SupF, confirming that the tnpR coding sequence specifies a single protein having both functions.