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Transcriptional analysis of pqqD and study of the regulation of pyrroloquinoline quinone biosynthesis in Methylobacterium extorquens AM1

Ramamoorthi, Roopa and Lidstrom, Mary E. (1995) Transcriptional analysis of pqqD and study of the regulation of pyrroloquinoline quinone biosynthesis in Methylobacterium extorquens AM1. Journal of Bacteriology, 177 (1). pp. 206-211. ISSN 0021-9193. http://resolver.caltech.edu/CaltechAUTHORS:RAMjbact95

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Abstract

Methanol dehydrogenase, the enzyme that oxidizes methanol to formaldehyde in gram-negative methylotrophs, contains the prosthetic group pyrroloquinoline quinone (PQQ). To begin to analyze how the synthesis of PQQ is coordinated with the production of other methanol dehydrogenase components, the transcription of one of the key PQQ synthesis genes has been studied. This gene (pqqD) encodes a 29-amino- acid peptide that is thought to be the precursor for PQQ biosynthesis. A unique transcription start site was mapped to a guanidine nucleotide 95 bp upstream of the pqqD initiator codon. RNA blot analysis identified two transcripts, a major one of 240 bases encoding pqqD and a minor one of 1,300 bases encoding pqqD and the gene immediately downstream, pqqG. Both transcripts are present at similar levels in cells grown on methanol and on succinate, but the levels of PQQ are about fivefold higher in cells grown on methanol than in cells grown on succinate. These results suggest that PQQ production is regulated at a level different from the transcription of pqqD. The genes mxbM, mxbD, mxcQ, mxcE, and mxaB are required for transcription of the genes encoding the methanol dehydrogenase subunits and were assessed for their role in PQQ production. PQQ levels were measured in mutants defective in each of these regulatory genes and compared with levels of pqqD transcription, measured with a transcriptional fusion between the pqqD promoter and xylE. The results showed that only a subset of these regulatory genes (mxbM, mxbD, and mxaB) is required for transcription of pqqD, and only mxbM and mxbD mutants affected the final levels of PQQ significantly.


Item Type:Article
Additional Information:Copyright © 1995, American Society for Microbiology. Received 14 July 1994/Accepted 28 October 1994. We thank Ludmila Chistoserdova for advice with the RNA experiments, Juan Davagnino for help with the ethanol dehydrogenase apoenzyme purification, and Francis Biville for help with the PQQ assays. This work was supported by grants from NIH (GM36296) and DOE (DEFG03-87ER13753).
Record Number:CaltechAUTHORS:RAMjbact95
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:RAMjbact95
Alternative URL:http://jb.asm.org/cgi/content/abstract/177/1/206
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ID Code:8813
Collection:CaltechAUTHORS
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Deposited On:19 Sep 2007
Last Modified:26 Dec 2012 09:42

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