The sigA gene encoding the major {sigma} factor of RNA polymerase from the marine cyanobacterium Synechococcus sp. strain PCC 7002: cloning and characterization

Department of Biochemistry and Molecular Biology, and Center for Biomolecular Structure and Function, The Pennsylvania State University, University Park, PA 16802, USA ABSTRACT Summary: The gene encoding the principal factor from Synechococcus sp. strain PCC 7002 was isolated and characterized. The Synechococcus sp. strain PCC 7002 sigA gene encodes a protein of 375 amino acids (43.7 kDa) that is required for viability under normal growth conditions. The SigA protein was overproduced in Escherichia coli and the purified protein was used to raise polyclonal antiserum in rabbits. This antiserum was used in immunoblot analyses of partially purified RNA polymerase from Synechococcus sp. strain PR6000. The probable in vivo translational start site was identified by a comparison of amino acid sequencing results obtained with SigA proteins overproduced in E. coli with immunoblot analyses of SigA protein in crude preparations of RNA polymerase from the cyanobacterium. The sigA gene is encoded on a transcript of 1700 bases that initiates 496 nucleotides upstream from the probable in vivo translational start site. The abundance of sigA transcripts decreases rapidly after the removal of combined nitrogen from the growth medium. Author for correspondence: Donald A. Bryant. Tel: +1 814 865 1992. Fax: +1 814 863 7024. e-mail: dab14@psuvm.psu.edu Keywords: Synechococcus sp., RNA polymerase, sigma factors, cyanobacteria Present address: Department of Microbiology, 527 Biological Sciences Building, University of Georgia, Athens, GA 30602-2605, USA.