Insertional Mutations in the Hydrogenase vhc and frc Operons Encoding Selenium-Free Hydrogenases in Methanococcus voltae.

Applied and Environmental Microbiology
Y Berghofer, A Klein

Abstract

Methanococcus voltae, which contains four different gene groups that encode [NiFe]-hydrogenases, was transformed with integration vectors to achieve polar inactivation of two of the four hydrogenase operons that encode the selenium-free enzymes Vhc and Frc. Transformants which were selected by their acquired puromycin resistance showed site-specific insertions in either the vhc or frc operon by single crossover events. Southern hybridization revealed tandem integrations of whole vectors in the vhc operon, whereas only one vector copy was found in the frc operon. Northern (RNA) hybridizations showed a pac transcript of defined size, indicating strong termination in front of the hydrogenase genes downstream. In spite of the apparent abolition of expression of selenium-free hydrogenases through these polar insertions, they were not lethal to cells upon growth in selenium-deprived minimal medium, which we had previously shown to strongly induce transcription of the respective operons in M. voltae. Instead, like wild-type control cultures, transformants responded to selenium deprivation only with a reduction in growth rate. We conclude that loss of the potential to express a selenium-free hydrogenase can nevertheless be balanced by ...Continue Reading

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Citations

Jul 20, 1999·Molecular Microbiology·D L Tumbula, W B Whitman
Jan 5, 2005·Nature Reviews. Genetics·Thorsten Allers, Moshe Mevarech
Jun 29, 2012·The ISME Journal·Christopher B WalkerDavid A Stahl
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Jun 14, 2001·Systematic and Applied Microbiology·Y Luo, A Wasserfallen
Apr 7, 2009·Biochimica Et Biophysica Acta·Tilmann Stock, Michael Rother

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