Multiple copies of the insertion-DNA sequences IS1 and IS2 in the chromosome of E. coli K-12

Saedler, Heinz; Heiss, Brian

doi:10.1007/bf00278602

Cited by 137 publications

(41 citation statements)

References 25 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…2A, lane 1, and 2B, lane 5). 35 S-labeled proteins were not produced by these cells without IPTG induction in the presence of rifampin (Fig. 2A, lane 3, and 2B, lane 7), showing that rifampin effectively suppressed host protein synthesis.…”

Section: Detection Of a 46-kda Is2 Proteinmentioning

confidence: 95%

See 1 more Smart Citation

Detection of an IS2-encoded 46-kilodalton protein capable of binding terminal repeats of IS2

Hu¹,

Lee²,

Lei³

1996

J Bacteriol

View full text Add to dashboard Cite

Insertion sequence IS2 is a small transposable element which is present in multiple copies in Escherichia coli K-12 and B strains as well as in other enteric bacteria (13,22,28,35). In addition to being a common cause of polar effects (9, 12), IS2 is also responsible for the transposition of the STII enterotoxin gene of enterotoxigenic E. coli (15,23). Nucleotide sequences of IS2 from three different sources have been reported (11,15,34). These three sequences are essentially identical. The entire IS2 is composed of 1,331 bp, which contains a pair of imperfect repeats of 42 bp at the terminus. In the original assignment of nucleotide numbers the IS2 sequence was counted from right to left (11,34). The terminal repeat located on the left-hand end of IS2 was called the right terminal inverted repeat (RIR), and that on the right-hand end was called the left terminal inverted repeat (LIR). These designations have created much confusion and inconvenience. In this report, the coordinates of IS2 are reoriented and the nucleotide positions are renumbered (Fig. 1). For comparisons, the original nucleotide numbers are shown in brackets immediately following the new numbers throughout the text and in the figures.Although the genome of IS2 contains five open reading frames (ORFs) which are capable of encoding proteins greater than 50 amino acids, only one, which has a molecular weight of 14,000, had been detected (14). This protein, InsA (14) A protein of 46 kDa was predicted to be produced via a Ϫ1 frameshift mechanism (7,8,16,30,(36)(37)(38)(39)42). In this communication, we report the detection of this 46-kDa protein of IS2. We also show that this 46-kDa protein can bind both terminal inverted repeats of IS2. MATERIALS AND METHODSRecombinant DNA techniques and materials. Plasmid DNA was isolated by the alkaline lysis method of Birnboim and Doly (3) and purified with a CsClethidium bromide density gradient. Recombinant DNA techniques were performed according to the methods described by Maniatis et al. (26). Restriction enzymes, the E. coli DNA polymerase I Klenow fragment, T4 polynucleotide kinase, and T4 DNA ligase were purchased from Boehringer Mannheim Biochemicals (Indianapolis, Ind.). Chemicals used in this study were obtained from Sigma Chemical Co. (St. Louis, Mo.) or Merck (Darmstadt, Germany).Analysis of IS2-encoded proteins. E. coli JM109(DE3) cells (40) containing pT7insA ( Fig. 2A), pT7insABЈ(A6) (Fig. 2B), or pETinsABЈ(A7) (Fig. 2C) were grown overnight at 37ЊC in 2 ml of M9 medium (26) containing 50 g of ampicillin per ml. This overnight culture was mixed with 2 ml of fresh M9 medium and was further incubated at 37ЊC with shaking until the A 600 of the culture reached 0.4. The cells in the culture were pelleted, washed with M9 buffer (26), and suspended in 1 ml of a methionine assay medium (methionine free) (Difco Laboratories, Detroit, Mich.). After an incubation at 37ЊC for 100 min, isopropyl-␤-D-thiogalactopyranoside (IPTG) was added to the culture to a final concentration of 1 mM to induce the production...

show abstract

Section: Detection Of a 46-kda Is2 Proteinmentioning

confidence: 95%

“…

…”

mentioning

confidence: 99%

Detection of an IS2-encoded 46-kilodalton protein capable of binding terminal repeats of IS2

Hu¹,

Lee²,

Lei³

1996

J Bacteriol

View full text Add to dashboard Cite

show abstract

“…ISI is present in various copy numbers in chromosomes as well as plasmids of certain bacteria belonging to the family Enterobacteriaceae (6)(7)(8)(9). ISl from plasmid R100 is 768 base pairs (bp) long (10); IS] elements from the other sources have divergent sequences (7,11,12) and hence are collectively termed iso-IS) elements (7).…”

mentioning

confidence: 99%

Frameshifting is required for production of the transposase encoded by insertion sequence 1.

Sekine

Ohtsubo

1989

Proc. Natl. Acad. Sci. U.S.A.

122

113

View full text Add to dashboard Cite

Insertion sequence IS] has two coding frames, insA and insB, which are essential for its transposition. Here, we show that a frameshifting event in the -1 direction from the 3' end region of the insA frame to an open reading frame (B' frame), extending from the 5' end of the insB frame, is involved in production of the InsA-B'-InsB fusion protein that has IS) transposase activity. The frameshifting event is likely to have occurred at the sequence AAAAAC where the insA frame overlaps the B' frame. Interestingly, this sequence is also present in one of the two sequences identified in retroviruses as frameshift signals for production of transframe polyproteins from the overlapping genes gag-pro or gag-propol.

show abstract

“…33 Small (typicallyo2 kb) insertion sequence (IS) elements were discovered a few years later as causes of spontaneous mutation in E. coli. [34][35][36] Most IS elements contain only genes and sites needed for their own regulated transposition, and like Mu, were intriguing as likely microbial counterparts of McClintock's maize elements. Superimposed on Mu's highly efficient transposition during lytic growth and during lysogenization were many features that seemed specific to its distinctive temperate phage life style.…”

Section: Immediate Implications Of Kan Insert Mapping and Early Tranmentioning

confidence: 99%

Julian Davies and the discovery of kanamycin resistance transposon Tn5

Berg

2016

J Antibiot

View full text Add to dashboard Cite

This paper recounts some of my fond memories of a collaboration between Julian Davies and myself that started in 1974 in Geneva and that led to our serendipitous discovery of the bacterial kanamycin resistance transposon Tn5, and aspects of the lasting positive impact of our interaction and discovery on me and the community. Tn5 was one of the first antibiotic resistance transposons to be found. Its analysis over the ensuing decades provided valuable insights into mechanisms and control of transposition, and led to its use as a much-valued tool in diverse areas of molecular genetics, as also will be discussed here. The Journal of Antibiotics (2017) 70, 339-346; doi:10.1038/ja.2016.120; published online 12 October 2016 BACKGROUND It was wonderfully educational, productive and exciting for me to work with Julian Davies for a few years in the mid 1970s, doing experiments that led to the serendipitous discovery and initial characterizations of kanamycin resistance transposon Tn5 1 -one of the first and most useful of the many bacterial antibiotic resistance (AR) transposons that have been found. 2 I was inspired by Julian's knowledge of mechanisms of antibiotic action and resistance mechanisms, his creativity and clarity of thought and expression, as illustrated in his numerous papers, 3-6 and by his high energy, kindness and inventive good humor-traits that have enriched the lives and careers of many people over the years. Described here, in tribute to Julian and his special character, are my memories of a collaboration that led to Tn5's unexpected discovery, Tn5's properties, and some lasting impacts of our collaboration on the field and on me personally.We began interacting in 1974, soon after Julian arrived in Geneva to begin his sabbatical with Pierre Francois Spahr (Department of Molecular Biology of the University). 7 I was then in my fourth (final) year as Chargé de Recherche (senior postdoc) in Lucien Caro's group in the same department, studying λdv plasmids, often in collaboration with Grete Kellenberger-Gujer, also in Lucien's group 8-13 -and during these years, also hugely enjoying features in and out of the lab that have drawn researchers to Geneva for decades: a superb scientific atmosphere, year-round easy access to the exquisite Alps and Jura mountains, and the language, cuisine and culture of Geneva and vicinity. I had become intrigued the year before by Julian's ideas about origins and evolution of resistance genes, as described in his paper with Raoul Benveniste. 14 They advanced the then-novel theory that the types of bacteria that had provided life saving antibiotics were also the ancestral sources of AR genes that were becoming increasingly common in human and veterinary pathogens. Their inferences were

show abstract

Multiple copies of the insertion-DNA sequences IS1 and IS2 in the chromosome of E. coli K-12

Cited by 137 publications

References 25 publications

Detection of an IS2-encoded 46-kilodalton protein capable of binding terminal repeats of IS2

Detection of an IS2-encoded 46-kilodalton protein capable of binding terminal repeats of IS2

Frameshifting is required for production of the transposase encoded by insertion sequence 1.

Julian Davies and the discovery of kanamycin resistance transposon Tn5

Contact Info

Product

Resources

About