LexA regulates the bidirectional hydrogenase in the cyanobacterium Synechocystis sp. PCC 6803 as a transcription activator

Gutekunst, Kirstin; Phunpruch, Saranya; Schwarz, Christoph; Schuchardt, Sven; Schulz-Friedrich, Rüdiger; Appel, Jens

doi:10.1111/j.1365-2958.2005.04867.x

LexA regulates the bidirectional hydrogenase in the cyanobacterium Synechocystis sp. PCC 6803 as a transcription activator

Author(s)

Gutekunst, Kirstin

Phunpruch, Saranya

Schwarz, Christoph

Schuchardt, Sven

Schulz-Friedrich, Rüdiger

Appel, Jens

Date Issued

November 1, 2005

Type

Article

DOI

10.1111/j.1365-2958.2005.04867.x

Abstract

The bidirectional NiFe-hydrogenase of Synechocystis sc. PCC 6803 is encoded by five genes (hoxEFUYH) which are transcribed as one unit. The transcription of the hox-operon is regulated by a promoter situated upstream of hoxE. The transcription start point was located at -168 by 5′Race. Several promoter probe vectors carrying different promoter fragments revealed two regions to be essential for the promoter activity. One is situated in the untranslated 5′leader region and the other is found -569 to -690 nucleotides upstream of the ATG. The region further upstream was shown to bind a protein. Even though an imperfect NtcA binding site was identified, NtcA did not bind to this region. The protein binding to the DNA was purified and found to be LexA by MALDI-TOF. The complete LexA and its DNA binding domain were overexpressed in Escherichia coli. Both were able to bind to two sites in the examined region in band-shift-assays. Accordingly, the hydrogenase activity of a LexA-depleted mutant was reduced. This is the first report on LexA acting not as a repressor but as a transcriptional activator. Furthermore, LexA is the first transcription factor identified so far for the expression of bidirectional hydrogenases in cyanobacteria. © 2005 The Authors. Journal compilation © 2005 Blackwell Publishing Ltd.

Citation

Molecular Microbiology, 58(3), 810-823, 2005