Berlin, Freie Universiẗat, Diss., 2001. / Dateiformat: zip, Dateien im PDF-Format.

Polyhydroxyalkanoate (PHA) is a cellular storage polymer with similar properties to some plastics when extracted from the cell and acts as a renewable and biodegradable plastic alternative. The objective of this research was to identify thermophilic PHA producing microorganisms and characterize their PHA accumulation, by which the process may become more competitive to current petroleum-based plastic production. This study characterizes growth and PHA production of Thermus thermophilus HB8 and three strains of T. filiformis (MOK, NGM and WRT), while simultaneously performing a genetic analysis for the presence of a phaC gene that encodes the PHA synthase. The results from the genetic analyses, together with attempts to produce PHA in T. thermophilus HB8 and T. filiformis strain (WRT, NGM and MOK) infer an absence of PHA accumulation. Contrary to our expectations, Thermus sp. is not a strong candidate for the thermophilic production of PHA.

Polyhydroxyalkanoate (PHA) is a cellular storage polymer with similar properties to some plastics when extracted from the cell and acts as a renewable and biodegradable plastic alternative. The objective of this research was to identify thermophilic PHA producing microorganisms and characterize their PHA accumulation, by which the process may become more competitive to current petroleum-based plastic production. This study characterizes growth and PHA production of Thermus thermophilus HB8 and three strains of T. filiformis (MOK, NGM and WRT), while simultaneously performing a genetic analysis for the presence of a phaC gene that encodes the PHA synthase. The results from the genetic analyses, together with attempts to produce PHA in T. thermophilus HB8 and T. filiformis strain (WRT, NGM and MOK) infer an absence of PHA accumulation. Contrary to our expectations, Thermus sp. is not a strong candidate for the thermophilic production of PHA.

Characterization of allosteric properties of phosphofructokinase from the extreme thermophile Thermus thermophilus (TtPFK) using thermodynamic linkage analysis revealed several peculiarities. Inhibition and activation of Fru-6-P binding by the allosteric effectors phosphoenolpyruvate (PEP) and MgADP are entropically-driven in TtPFK. It is also curious that PEP binding affinity is unusually strong in TtPFK when compared to PFKs from Escherichia coli, Bacillus stearothermophilus, and Lactobacillus delbrueckii, while the magnitude of the allosteric inhibition by PEP is much smaller in TtPFK. In an effort to understand the source of weak inhibition, a putative network of residues between the allosteric site and the nearest active site was identified from the three-dimensional structures of BsPFK. Three of the residues in this network, D59, T158, and H215, are not conserved in TtPFK, and, due to their nature (N59, A158, S215), are unlikely to be involved in the same non-covalent interactions seen in BsPFK. The triple chimeric substitution N59D/A158T/S215H, results in a 2.5 kcal mol-1 increase in the coupling free energy, suggesting that the region containing these residues may be important for propagation of inhibitory response. The individual substitutions at each position resulted in an increase in the coupling free energy, and the double substitutions displayed additivity of these changes. The chimeric substitution made at N59 suggests that the polar nature of the asparagine at position 59 is key for the enhanced binding of PEP. The non-conserved R55 was found to be particularly important for the enhanced binding of PEP in TtPFK, as chimeric substitutions R55G and R55E resulted in a 3.5 kcal mol-1 and 4.5 kcal mol-1 decrease in the binding affinity for PEP, respectively. Our results also confirm the observations previously made in PFKs from E. coli and B. stearothermophilus, that the ability of the effector to bind is independent of its ability to produce allosteric response. We show that several substitutions result in a decrease in binding affinity of PEP to TtPFK, while dramatically enhancing its ability to inhibit (N59D, R55G, R55E). Similarly, some substitutions, like S215H and A158T show an enhanced inhibition by PEP, while having no effect on its binding affinity.

Frankfurt (Main), Universiẗat, Diss., 2008. / Erscheinungsjahr an der Haupttitelstelle: 2007.

Frankfurt (Main), Univ., Diss., 2004. / Erscheinungsjahr an der Haupttitelstelle: 2003. Computerdatei im Fernzugriff.

Bielefeld, Univ., Diss., 2001. / Computerdatei im Fernzugriff.

Bielefeld, Univ., Diss., 2001. / Computerdatei im Fernzugriff.

Bielefeld, University, Diss., 2001.

Frankfurt (Main), Universiẗat, Diss., 2004. / Erscheinungsjahr an der Haupttitelstelle: 2003.