Control analysis of mixed populations of gluconobacter oxydans and saccharomyces cerevisiae

[摘要] ENGLISH ABSTRACT:In the last decade a need arose to find a theoretical framework capable of gaining a quantitativeunderstanding of ecosystems. Control analysis was proposed as a suitable candidate for the analysisof 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 throughexperimental means by applying techniques akin to enzyme kinetics of biochemistry on a simpleecosystem between Saccharomyces cerevisiae and Gluconobacter oxydans. However, this exercisewas far more complex than we originally expected due to the extra metabolic activities presented byboth organisms.Nevertheless, we derived suitable kinetic equations to describe the metabolic behaviour of bothorganisms, with regards to the activities of interest to us, from pure culture experiments. Wedeveloped new techniques to determine ethanol and oxygen sensitivity of G. oxydans based on itsobligately aerobic nature. These parameters were then used to build a simple kinetic model and amore complex model incorporating oxygen limited metabolism we observed at higher cell densitiesof G. oxydans. Our models could predict both situations satisfactorily for pure cultures andespecially the more complex model could describe the lack of linearity observed between metabolicactivity 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 ethanolconcentration and acetate flux, which was a positive indication for the application of controlanalysis on the ecosystem. However, the theoretical models based on parameters derived from pureculture experiments did not predict mixed culture steady states accurately. Careful analysis showedthat 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 fromthe experimental data obtained from mixed cultures. We could calculate the control coefficientsdirectly from the experimental data of mixed population studies and compare it with those fromtheoretical models based on 3 different parameter sets. Our analysis showed that the yeast had allthe 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 inmind, 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 informationobtained especially regarding control structure with regards to the steady-state ethanolconcentration.

[发布机构] Stellenbosch University