Department of Biochemistry

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Browsing Department of Biochemistry by Subject "Acetobacter"

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    Control analysis of mixed populations of gluconobacter oxydans and saccharomyces cerevisiae
    (Stellenbosch : University of Stellenbosch, 2010-12) Malherbe, Christiaan Johannes; Snoep, Jacob Leendert ; Rohwer, Johann M.; University of Stellenbosch. Faculty of Science. Dept. of Biochemistry.
    ENGLISH ABSTRACT: In the last decade a need arose to find a theoretical framework capable of gaining a quantitative understanding of ecosystems. Control analysis was proposed as a suitable candidate for the analysis of ecosystems with various theoretical applications being developed, i.e. trophic control analysis (TCA) and ecological control analysis (ECA). We set out to test the latter approach through experimental means by applying techniques akin to enzyme kinetics of biochemistry on a simple ecosystem between Saccharomyces cerevisiae and Gluconobacter oxydans. However, this exercise was far more complex than we originally expected due to the extra metabolic activities presented by both organisms. Nevertheless, we derived suitable kinetic equations to describe the metabolic behaviour of both organisms, with regards to the activities of interest to us, from pure culture experiments. We developed new techniques to determine ethanol and oxygen sensitivity of G. oxydans based on its obligately aerobic nature. These parameters were then used to build a simple kinetic model and a more complex model incorporating oxygen limited metabolism we observed at higher cell densities of G. oxydans. Our models could predict both situations satisfactorily for pure cultures and especially the more complex model could describe the lack of linearity observed between metabolic activity and cell density at higher cell densities of G. oxydans. Mixed populations of S. cerevisiae and G. oxydans reached quasi-steady states in terms of ethanol concentration and acetate flux, which was a positive indication for the application of control analysis on the ecosystem. However, the theoretical models based on parameters derived from pure culture experiments did not predict mixed culture steady states accurately. Careful analysis showed that these parameters were mostly under-estimated for G. oxydans and overestimated for S. cerevisiae. Hence, we calculated the kinetic parameters for mixed population assays directly from the experimental data obtained from mixed cultures. We could calculate the control coefficients directly from the experimental data of mixed population studies and compare it with those from theoretical models based on 3 different parameter sets. Our analysis showed that the yeast had all the control over the acetate flux while control over the steady-state ethanol was shared. The strength of our approach lies in designing our experiments with a control analysis approach in mind, but we have also shown that even for simple ecosystems this approach is non-trivial. Despite the various experimental challenges, this approach was very rewarding due to the extra information obtained especially regarding control structure with regards to the steady-state ethanol concentration.