2. Introduction
In extreme conditions, certain microorganisms have evolved unique methods
necessary to adapt to high salt, pH, or temperature conditions. Of these
“extremophiles”, those in the Archaea domain of life have immense implications for
biotechnology. Understanding how these microorganisms adapt to extreme
conditions can help humans apply the same techniques in industry to develop
genetically engineered products. Many archaea have the potential for significant
biotechnological advantages, but have not yet been fully developed.
3. Current Literature
Glucose can offset defects arising from deletion of transcription factor trmB, which is
associated with metabolic pathways in Halobacterium salinarium (Schmid et al.,
2009).
Arabinate, a derivative of glucose, has not yet been tested with respect to cell
growth.
Studying arabinate can help specify the function of trmB and its corresponding
metabolic gene network while also developing use of the sugar in future studies of
halobacteria.
4. Hypotheses
Does introducing various concentrations of glucose compensate for the known
growth deficiency in the Δtrmb strain?
H0: μtrmb/glu = μ trmb
H1: μtrmb/glu > μ trmb
As a glucose derivative, does arabinate reproduce the growth trends found in the
Δtrmb strain with glucose?
H0: μtrmb/glu ≠ μ trmb/arab
H2: μtrmb/glu = μ trmb/arab
5. Serial Dilution of Sugars
*Note: After adding the sugar mixture to the cells at a 1:1 ratio, the above dilutions were halved
6. Methods
H. salinarum and a trmB knockout strain were collected
Both strains were diluted in growth media and treated with 5 concentrations of
glucose or arabinate.
We performed a 1:5 serial dilution from 10% and added a control group.
Following 1.5 days of incubation with the optical densities analyzed every 30
minutes, the archaea and data were collected.
7. The following graphs display the logistically fitted
growth rates of H. salinarum at four concentrations of
arabinate or glucose solution and a sugarless control.
8. Figure A: Serves to demonstrate that the wild type strain of the archaea is not as
affected by the addition of sugar as the trmB-deficient mutant. Demonstrated by
the small difference between the lines steady states.
9. Figure B: Demonstrates that adding glucose will still affect the growth rate of the
archaea, similar to the way archaea responded to arabinate. Could be a result of
lab error and requires further examination.
10. Figure C: Suggests that 1% arabinate is the optimum concentration for
maximizing growth rate in the absence of the gene trmB.
11. Figure D: Introduction of glucose into mutant H. salinarum compensates for the
lack of trmB. Limit to the amount of glucose that can be added beneficially,
between concentrations of 0.20% and 0.04%.
12. Figure E: Compares the growth rates of the mutant strain ΔtrmB of H. salinarum
in 5% sugar concentration with both ΔtrmB in sugarless control and Δura3 in 0%
sugar. Simultaneously demonstrates the effect of adding sugar to ΔtrmB to
induce increased growth rate and compares that result with the wild type still
with the trmB gene.
Arabinate Glucose
Sugar
MaximumGrowthRate(cells/mL/hour)
13. Quick Results
While testing different solution concentrations with each strain of archaea, we found
a correlation between increased growth rate and increased sugar concentration.
The control group of ΔtrmB had a maximum growth lower than that of the wild type
control group.
14. Key Points from Figures
When glucose is added to the ΔtrmB mutant, the growth rate increases (Fig. D).
Similarly, with arabinate, the original growth rate of the mutant archaea started out
lower than the wild type (Fig. E).
With the addition of 5% arabinate solution, there was a 29.4% increase in the growth
rate of ΔtrmB and a 28.6% increase in the growth rate of Δura3.
While in previous glucose studies was found to have a negligible impact on growth of
wild type archaea (Schmid et.al., 2009), we found that arabinate improved growth in
the wild type (Fig. E ).
15. Further Research
It is possible to test the growth of H. salinarum in other different types of sugar, such
as ribose, to see if it responds in a similar manner
We can fine tune the optimal sugar concentration by performing smaller iterations in
the series. Research into which sugar improve H. salinarum growth furthers our
knowledge about the metabolic pathways of them and their potential uses.
16. Acknowledgements and Citations
Thanks to the following individuals for contributing to our project:
Allison Edgar
Jordan Gulli
Paul Magwene
Colin Maxwell
Amy Schmid
Rotem Ben-Shachar
Amy Sheck
Horia Todor
Peter Tonner
Jennifer Wygoda
Works Cited
Schmid, A. K., David, R. J., Pan, M., Kolde, T., & Baliage, N. S. (2009). A single transcription factor regulates evolutionarily
diverse but functionally linked metabolic pathways in response to nutrient availability. Molecular Systems Biology,
5(282), 1-15. doi: 10.1038/msb.2009.40.
