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Mechanistic Characterization of Transaldolase from Thermoplasma Acidophilum and Preliminary Analysis of the QncN/L-M Protein System from Streptomyces Melanovinaceus

dc.contributor.advisorTittmann, Kai Prof. Dr.
dc.contributor.authorSautner, Viktor
dc.date.accessioned2018-02-13T09:55:30Z
dc.date.available2018-02-13T09:55:30Z
dc.date.issued2018-02-13
dc.identifier.urihttp://hdl.handle.net/11858/00-1735-0000-002E-E350-8
dc.identifier.urihttp://dx.doi.org/10.53846/goediss-6713
dc.language.isoengde
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/
dc.subject.ddc572de
dc.titleMechanistic Characterization of Transaldolase from <i>Thermoplasma Acidophilum</i> and Preliminary Analysis of the QncN/L-M Protein System from <i>Streptomyces Melanovinaceus</i>de
dc.typedoctoralThesisde
dc.contributor.refereeTittmann, Kai Prof. Dr.
dc.date.examination2016-08-23
dc.description.abstractengThe results introduced in the present work are mainly based on the pre-steady state and steady-state analysis supported by structural information from crystallographic studies. Different variants of TacTAL were generated and analyzed, using these methods. The “closed” conformation of TacTAL could be arrested in the TacTALT30C/D211C variant. It could be shown that the „closed“ state is important for the binding of the substrate F6P and the catalysis of the donor half-reaction. On the other hand, the “open” conformation is necessary for the acceptor half-reaction. Furthermore, the transaldolase activity could be transformed into fructose-6-phosphate aldolase activity of the TacTALE60Q/F132Y variant. The acid-base situation in the active site of the variant is the same as in the active site of natural EcFSA. It could be shown, that the introduced Tyr132 residue takes over the role of the general acid-base catalyst Glu60. This tyrosine residue is placed in favorable orientation for the direct protonation of the central carbanion/enamine intermediate, which discriminates between the aldolase and transaldolase activity. The analysis of the interaction between the substrate’s C1 hydroxy group and the active site showed that the deletion of the respective interaction partner on the part of the enzyme (TacTALN108A/S130A variant) or on the part of the substrate (A5P as substrate) results in comparable decrease of the donor half-reaction activity. Finally, the binding mode of the substrate F6P in the active site of TacTAL as Michaelis complex was analyzed in the structure of the catalytically inactive variant TacTALK86Q co-crystallized with F6P. The relative orientation of the F6P-carbinolamine intermediate in the active site was analyzed in the structure of TacTALwt co-crystallized with the carbinolamine mimic M1P. Additionally to the studies on transaldolase from Thermoplasma acidophilum, the protein system containing the QncN/L, QncM from Streptomyces melanovinaceus and the E3 component from Streptomyces lividans was preliminarily characterized.de
dc.contributor.coRefereeRodnina, Marina Prof. Dr.
dc.subject.engTransaldolasede
dc.subject.engAldolasede
dc.subject.engThermoplasmade
dc.subject.engAcidophilumde
dc.subject.engKineticsde
dc.subject.engStructurede
dc.identifier.urnurn:nbn:de:gbv:7-11858/00-1735-0000-002E-E350-8-8
dc.affiliation.instituteGöttinger Graduiertenschule für Neurowissenschaften, Biophysik und molekulare Biowissenschaften (GGNB)de
dc.subject.gokfullBiologie (PPN619462639)de
dc.identifier.ppn1013847768


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