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Biophysical characterization of heterocyst differentiation regulators, HetR and PatS, from the cyanobacterium, Anabaena sp. strain PCC 7120 and structural biology of bacterial proteins from the Northeast Structural Genomics Consortium

Feldmann, Erik A.
http://rave.ohiolink.edu/etdc/view?acc_num=miami1342801532

Abstract Details

2012, Doctor of Philosophy, Miami University, Chemistry and Biochemistry.
The filamentous cyanobacterium Anabaena (Nostoc) sp. strain PCC 7120 has evolved a mechanism for isolating the two incompatible processes of photosynthesis and nitrogen fixation by differentiating specialized cells called heterocysts. The structure and metabolism of heterocysts support a microanaerobic environment allowing for the nitrogenase-dependent fixation of dinitrogen into ammonia to occur with limited inhibition by oxygen. Under conditions of nitrogen starvation, Anabaena differentiates heterocysts in regular patterned intervals in approximately one out of every ten cells. The hetR and patS genes are essential for proper heterocyst differentiation. The gene products, HetR and PatS, are regulators of heterocyst differentiation, but their specific mechanisms of activity are unclear. In the first part of this dissertation, an in depth biophysical characterization of both HetR and PatS is presented. We describe a method for over-expressing and purifying high yields of soluble HetR from Anabaena in Escherichia coli, have provided the first experimental evidence that the master regulatory HetR transcription factor is the direct binding target for the PatS inhibitor peptide, have characterized the binding stoichiometry of HetR with DNA and PatS substrates, and have quantified the binding thermodynamics of HetR-PatS complexes. The second part of this dissertation describes a detailed structural biology summary of four different bacterial protein targets of the National Institutes of Health. The first section of this second part includes an uncharacterized protein, Pspto_3016 from Pseudomonas syringae which adopts a putative DNA-binding double-wing motif. The second and third sections include two different proteins from the cyanobacterium Anabaena sp. PCC 7120: a hypothetical uncharacterized protein, Asl3597, with a novel fold and that exhibits sequence homology to the chlororespiratory reductase family of plant proteins, and another uncharacterized protein, Asr4154, annotated as a subunit of the protochlorophyllide reductase superfamily of enzymes. The fourth and final section includes an uncharacterized protein fragment, the putative mucin-binding domain of the protein LBA1460 from Lactobacillus acidophilus. We provide high resolution, three-dimensional structures of each protein, solved by solution NMR spectroscopy, to assist in future characterization of their corresponding biological and biochemical functions.
Michael A. Kennedy, PhD (Advisor)
Christopher A. Makaroff, PhD (Committee Chair)
Dabney-Smith Carole, PhD (Committee Member)
Michael W. Crowder, PhD (Committee Member)
Richard E. Lee, PhD (Committee Member)
194 p.
Biochemistry; Biophysics; Chemistry; Developmental Biology
Anabaena PCC 7120; Nostoc; HetR; PatS; heterocyst differentiation; structural genomics; NESG; EPR spectroscopy; ITC calorimetry; multidimensional NMR spectroscopy; protein NMR

Recommended Citations

Citations

  • Feldmann, E. A. (2012). Biophysical characterization of heterocyst differentiation regulators, HetR and PatS, from the cyanobacterium, Anabaena sp. strain PCC 7120 and structural biology of bacterial proteins from the Northeast Structural Genomics Consortium [Doctoral dissertation, Miami University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=miami1342801532

    APA Style (7th edition)

  • Feldmann, Erik. Biophysical characterization of heterocyst differentiation regulators, HetR and PatS, from the cyanobacterium, Anabaena sp. strain PCC 7120 and structural biology of bacterial proteins from the Northeast Structural Genomics Consortium. 2012. Miami University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=miami1342801532.

    MLA Style (8th edition)

  • Feldmann, Erik. "Biophysical characterization of heterocyst differentiation regulators, HetR and PatS, from the cyanobacterium, Anabaena sp. strain PCC 7120 and structural biology of bacterial proteins from the Northeast Structural Genomics Consortium." Doctoral dissertation, Miami University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=miami1342801532

    Chicago Manual of Style (17th edition)

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© 2012, all rights reserved.
This open access ETD is published by Miami University and OhioLINK.