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Nucleic Acids Research, 1990, Vol. 18, No. 3 619-624
© 1990

MOLECULAR BIOLOGY

Identification of the metalloregulatory element of the plasmid-encoded arsenical resistance operon

Michael J.D.San Francisco+, Constance L. Hope, Joshua B. Owolabi, Louis S. Tisa§ and Barry P. Rosen*

* To whom correspondence should be addressed at Department of Biochemistry, Wayne State University, School of Medicine, Scott Hall of Basic Medical Sciences, 540 East Canfield, Detroit, MI 48201, USA

Received September 13, 1989. Revised December 18, 1989. Accepted December 18, 1989.

The regulatory region of the plasmid-encoded arsenical resistance (ars) operon was cloned as a 727-bp EcoRI-HindIII fragment. When cloned into a promoter probe vector this fragment conferred arsenite inducible tetracycline resistance in Escherichia coli, indicating that the fragment carried a regulatory gene, the arsR gene. A single region corresponding to –35 and –10 promoter recognition sites was identified. The transcriptional start site of the mRNA was determined by primer extension. The sequence has an open reading frame for a potential 13,179 Da polypeptide, termed the ArsR protein. The fragment was cloned into a temperature regulated expression vector. A protein with an apparent molecular mass of about 12 kDa was induced by either temperature or arsenite. This protein was purified and used to produce antibodies specific for the ArsR protein.

+Present addresses: Biotechnology Center, Rightmire Hall, Ohio State University Columbus, OH 43210, USA

§Department of Biochemistry, College of Agriculture and Life Sciences, 420 Henry Mall, University of Wisconsin Madison, Wl 53706, USA


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