Denitrification and the hypoxic response in obligate aerobic methane-oxidizing bacteria

• Author / Creator

  Kits, Kerim D

• Aerobic methanotrophic bacteria lessen the impact of the greenhouse gas methane (CH4) not only because they are a sink for atmospheric methane but also because they oxidize it before it is emitted to the atmospheric reservoir. Aerobic methanotrophs, unlike anaerobic methane oxidizing archaea, have a dual need for molecular oxygen (O2) for respiration and CH4 oxidation. Nevertheless, methanotrophs are highly abundant and active in environments that are extremely hypoxic and even anaerobic. While the O2 requirement in these organisms for CH4 oxidation is inflexible, recent genome sequencing efforts have uncovered the presence of putative denitrification genes in many aerobic methanotrophs. Being able to use two different terminal electron acceptors – hybrid respiration – would be massively advantageous to aerobic methanotrophs as it would allow them to halve their O2 requirement. But, the function of these genes that hint at an undiscovered respiratory anaerobic metabolism is unknown. Moreover, past work on pure cultures of aerobic methanotrophs ruled out the possibility that these organisms denitrify. An organism that can couple CH4 oxidation to NO3- respiration so far does not exist in pure culture. So while the role of aerobic methanotrophs in the carbon cycle is appreciated, the hypoxic metabolism and contribution of these specialized microorganisms to the nitrogen cycle is not understood. Here we demonstrate using cultivation dependent approaches, microrespirometry, and whole genome, transcriptome, and proteome analysis that an aerobic methanotroph – Methylomonas denitrificans FJG1 – couples CH4 oxidation to NO3- respiration with N2O as the terminal product via the intermediates NO2- and NO. Whole transcriptome and proteome analysis reveals that respiratory nitrate (Nar, Nap), nitrite (Nir), and nitric oxide (Nor) reductases are expressed and upregulated at the transcript and protein levels under denitrifying conditions. Physiological analysis of denitrifying cultures of M. denitrificans FJG1 also confirms that NO3- respiration is bioenergetically advantageous. We also confirm denitrification activity and upregulation of denitrification gene expression in another obligate methanotroph – Methylomicrobium album BG8 – and also show that this activity is supported by a diverse array of energy sources. Formaldehyde fermentation has been identified by others as an important adaptation of aerobic methanotrophs to hypoxia. We additionally illustrate here that M. denitrificans FJG1 can also ferment formaldehyde and that nitrate respiration and fermentation occur simultaneously during hypoxia. The work herein is a significant contribution to our knowledge of how aerobic methanotrophs affect greenhouse gas flux though CH4 oxidation in low O2 environments and through N2O production via denitrification.

• Subjects / Keywords

  • Methane
  • Nitrous oxide
  • Nitrite reduction
  • Denitrification
  • Nitrate respiration
  • Methylomicrobium
  • Methylomonas
  • Hypoxia
  • Methanotroph

• Graduation date

  Fall 2016

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/R3KP7V53Q

• License

  This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.