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Journal of Bacteriology, August 1998, p. 3900-3906, Vol. 180, No. 15
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
Activation Control of pur Gene Expression in
Lactococcus lactis: Proposal for a Consensus Activator
Binding Sequence Based on Deletion Analysis and Site-Directed
Mutagenesis of purC and purD Promoter
Regions
Mogens
Kilstrup,1,*
Stine G.
Jessing,1
Stephanie B.
Wichmand-Jørgensen,1
Mette
Madsen,2 and
Dan
Nilsson2
Department of Microbiology, Technical
University of Denmark, DK2800 Lyngby,1 and
Department of Physiology and Metabolism, Chr. Hansen A/S,
DK2870 Hørsham,2 Denmark
Received 23 March 1998/Accepted 28 May 1998
A comparison of the purC and purD upstream
regions from Lactococcus lactis revealed the presence of a
conserved ACCGAACAAT decanucleotide sequence located
precisely between
79 and
70 nucleotides upstream from the
transcriptional start sites. Both promoters have well-defined
10
regions but lack sequences resembling
35 regions for
70 promoters. Fusion studies indicated the importance of
the conserved sequence in purine-mediated regulation. Adjacent to the
conserved sequence in purC is a second and similar region
required for high-level expression of the gene. A consensus PurBox
sequence (AWWWCCGAACWWT) could be proposed for the three
regions. By site-directed mutagenesis we found that mutation of the
central G in the PurBox sequence to C resulted in low levels of
transcription and the loss of purine-mediated regulation at the
purC and purD promoters. Deletion analysis also showed that the nucleotides before the central CCGAAC core
in the PurBox sequence are important. All results support the idea that
purC and purD transcription is regulated by a
transcriptional activator binding to the PurBox sequence.
*
Corresponding author. Mailing address: Department of
Microbiology, Technical University of Denmark, Building 301, DK2800
Lyngby, Denmark. Phone: 45 45 25 25 28. Fax: 45 45 88 26 60. E-mail: mk{at}im.dtu.dk.
Journal of Bacteriology, August 1998, p. 3900-3906, Vol. 180, No. 15
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
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