The amplifiable unit of DNA no. 1 (AUD1) of Streptomyces lividans consists of three 1 kb repeats (left direct repeat, LDR; middle direct repeat, MDR; and the slightly different right direct repeat, RDR) and two 4.7 kb repeats alternately arranged in identical orientation to each other. Both 4.7 kb repeats have been sequenced. They are identical and contain one open reading frame (orf4.7). The deduced amino acid sequence has a low similarity to chitinases, and two amino acid repeats present high similarities to fibronectin type III modules. Sequencing had previously shown that the ORF corresponding to each 1 kb repeat encodes a putative DNA-binding protein. Crude extracts of Escherichia coli overexpressing the orfRDR-encoded protein and of S. lividans Jni1, having a high amplification of AUD1 and therefore orfMDR, were used in gel retardation assays. The orfRDR- and probably the orfMDR-encoded proteins can bind to an imperfect palindromic sequence upstream from MDR and RDR and to another sequence downstream from RDR. An extrachromosomal DNA amplification system was constructed containing different combinations of the sequences composing AUD1. In mutants having a deletion of the chromosomal AUD1, the 4.7 kb repeats could be reduced in size, mutated or replaced by E. coli DNA without altering the ability to amplify when RDR was present. Therefore, the only function of the 4.7 kb repeats in amplification is to provide directly repeated DNA sequences. When RDR was lacking or mutated, no amplification was observed. This strongly suggests that the DNA-binding protein encoded by orfRDR is required for AUD1 amplification.

The unstable linear chromosome of Streptomyces lividans was circularized by homologous recombination and its terminal inverted repeats deleted. Strains with circularized chromosomes showed no obvious phenotypic disadvantages compared to the wild type. However, they segregated about 20 times more chloramphenicol-sensitive mutants than the wild type (24.3% vs. 1.4%), due to a higher incidence of large deletions. In addition, in all circularized chromosomes amplification of 30-60 kb fragments was observed at the new chromosomal junction, to levels of approximately 10 copies per chromosome. Arginine auxotrophs that arose spontaneously among the progeny of strains with a circularized chromosome showed high-copy-number amplification of the DNA element AUD1, as also seen in mutants of the wild type. These observations demonstrate that the circular form of the Streptomyces chromosome is more unstable than the linear one.

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The junctions of the Streptomyces lividans chromosomal terminal inverted repeats (TIRs) were isolated from cosmid clones as 6.2 kb PstI and 2.9 kb BamHI fragments, respectively. The fragments were completely sequenced. In each of the fragments just one open reading frame could be identified. One putative gene product showed significant similarities to a sensor and the other to a transcriptional regulator protein of prokaryotic two-component signalling systems. Next to one TIR numerous long direct repeats were found within a region of about 400 bp.