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A new insertion sequence IS1452 from Acetobacter pasteurianus

Department of Biotechnology, Division of Agriculture and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113, Japan

²Author for correspondence: Sueharu Horinouchi. Tel: +81 03 3812 2111 ext. 5123. Fax: +81 3 5802 2931.

ABSTRACT

A new insertion sequence element, IS1452, was found to be associated with inactivation of the alcohol dehydrogenase by insertion in the adhS gene encoding subunit III of the three-component membrane-bound alcohol dehydrogenase complex in Acetobacter pasteurianus. Cloning and sequencing analyses of the mutated subunit III gene locus revealed that IS 1452 was inserted at or near the ribosome-binding sequence of adhS. Analysis of transcription using the chloramphenicol acetyltransferase gene as the reporter indicated that IS1452 abolished transcription of adhS by separating its promoter from the subunit III structural gene. IS1452 was 1411 bp in length and had a terminal inverted repeat of 21 bp. IS1452 contained one long ORF of 416 amino acids rich in basic amino acids. This protein showed homology with a putative transposase, Tra1, of IS701 isolated from the cyanobacterium Calothrix species PCC 7601. Like IS701, IS7452 was found to generate a 4 bp direct repeat at the site of insertion upon transposition. The target site specificity was rather strict, and a CTA(A or G) sequence appeared to be preferentially recognized. Transposition of IS1452 was replicative, since it was accompanied by an increase in the copy number of IS1452. Several strains belonging to the genus Acetobacter also contained IS1452 at varying copy numbers from one to more than ten. These observations suggest that IS1452 is one of the insertion sequences that are responsible for genetic instability leading to deficiencies in various physiological properties in acetic acid bacteria.

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