(191aq) Systematic Carbon and Growth Analysis of a Promising Methanotroph Strain

Systematic carbon and growth analysis of a promising methanotroph strain

Kyle Stone, Q.
Peter He, and Jin Wang

Department of
Chemical Engineering, Auburn University, Auburn, AL, 36849, USA

Methanotrophs are
widespread bacteria that use methane as their sole carbon and energy source¹.
In doing so the organisms, are not only able to reduce the global warming
potential of natural gas leaks or effluent from coal ventilation, but also
fulfill the role of a steady biocatalyst utilizing the 376-490 trillion Btu
worth of biogas made in the US². Due to recent developments in
genetic studies, metabolic modeling, and downstream processing, these cells
have been studied for commercial applications, including the production of
single cell protein (SCP), lipids (for biodiesel), methanol, poly-β-hydroxybutyrate (PHB), as well as waste treatment or
bioremediation³. One strain in particular, Methylomicrobium buryatense
5GB1, has been identified as a promising biocatalyst because it is a fast
growing organisms with the ability to generate a variety of products including
lipids for biodiesel, organic acids, and ectoine. Since the organism is haloalkaliphilic the high pH and salt concentrations are
inhibitory to many overpowering contaminants, as well ^3-6.

In this work we
provide a systematic approach in analyzing the carbon use and growth patterns
of 5GB1 with an in house gas mixing system that safely creates a variety of
headspace conditions with methane, oxygen, and nitrogen. Of particular interest
are the yields of methane to biomass, to carbon dioxide, and to organic liquid
products in batch cultures with balanced, methane limited, or oxygen limited
headspaces. The approaches used accurately measure carbon products with an
overall balance of 95%-102%, taking into account CO₂ that becomes
far more soluble in alkaliphilic conditions. This
solubility studied, illustrated the requirement of an effective Henry’s
constant to properly characterize the gas-liquid equilibrium of CO₂ and
to have an accurate carbon balance.

The growth patterns
observed thus far with the systematic comparison suggest that oxygen
concentration amongst different oxygen:methane
ratios plays a significant role on the optimal growth rate of this organism.
Specifically, an excess of oxygen leads to a general decrease of growth rates
in the cultures tested. Additionally, gas uptake rates are evaluated to further
gauge when substrate limitation occurs and to use as an indicator of possible
metabolic pathway changes. Overall the study contributes to the much needed
knowledge base for successful methane fermentation strategies with this
promising methanotroph.

1)     
Conrad R. 2009. The global methane cycle: recent advances
in understanding the microbial processes involved. Environ. Microbiol. Reports 1:285–292.

2)     
USDA. Biogas Opportunities Roadmap: 2014.

3)     
Kalyuzhnaya
MG, Puri AW, Lidstrom ME.
2015. Metabolic engineering in methanotrophic bacteria. Metab.
Eng. 29:142–152.

4)     
Gilman
A, Laurens LM, Puri AW, Chu F, Pienkos
PT, Lidstrom ME. 2015. Bioreactor performance
parameters for an industrially-promising methanotroph Methylomicrobium
buryatense 5GB1. Microb.
cell factories 14.

5)     
Henard CA, Smith H, Dowe N, Kalyuzhnaya MG, Pienkos
PT, Guarnieri MT. 2016. Bioconversion of methane to
lactate by an obligate methanotrophic bacterium. Sci. reports 6.

6)     
Strong
PJ, Kalyuzhnaya M, Silverman J, Clarke WP. 2016. A methanotroph-based biorefinery: potential scenarios for generating multiple
products from a single fermentation. Bioresour.
Technol. 215.