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1Departments of Microbial Physiology, Faculty of Biology, Biocentrum Amsterdam, Vrije Universiteit, De Boelelaan 1087, NL-1081 HV, The Netherlands
2Departments of Molecular Microbiology, Faculty of Biology, Biocentrum Amsterdam, Vrije Universiteit, De Boelelaan 1087, NL-1081 HV, The Netherlands
3Microbiology Research Centre Holland, Hudsonstraat 68, 1057 SN Amsterdam, The Netherlands
ABSTRACT
The mau gene cluster of Paracoccus denitrificans constitutes 11 genes (10 are located in the transcriptional order mauFBEDACJGMN; the 11th, mauR, is located upstream and divergently transcribed from these genes) that encode a functional methylamine-oxidizing electron transport branch. The mauR gene encodes a LysR-type transcriptional activator essential for induction of the mau operon. In this study, the characteristics of that process were established. By using lacZ transcriptional fusions integrated into the genome of P. denitrificans, it was found that the expression of the mauR gene during growth on methylamine and/or succinate was not autoregulated, but proceeded at a low and constant level. The mauF promoter activity was shown to be controlled by MauR and a second transcriptional regulator. This activity was very high during growth on methylamine, low on succinate plus methylamine, and absent on succinate alone. MauR was overexpressed in Escherichia coli by using a T7 RNA polymerase expression system. Gel shift assays indicated that MauR binds to a 403 bp DNA fragment spanning the mauR-mauF promoter region. It is concluded from these results that the expression of the structural mau genes is dependent on MauR and its inducer, methylamine, as well as on another transcription factor. Both activators are required for high-level transcription from the mauF promoter. It is hypothesized that the two activators act synergistically to activate transcription: the effects of the two activators are not additive and either one alone activates the mauF promoter rather weakly.
*Author for correspondence: Rob J. M. van Spanning. Tel: +31 20 444 7179. Fax: +31 20 444 7229. e-mail: spanning@bio.vu.nl
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