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Computational Studies of Protein Folding Assistance and Conformational Pathways of Biological Nanomachines

Smith, Nathan B.

Abstract Details

2015, PhD, University of Cincinnati, Arts and Sciences: Chemistry.
Protein homeostasis is crucial for a cell's viability and therefore an organism's survival. Proteins, upon translation, must reach a three-dimensional structure known as their native state. This structure defines a particular protein’s function from a set of diverse enzymatic activities. Proteins must traverse a rough, many multi-dimensional energy landscape in order to approach their native state at the native basin of attraction. While some small, single-domain proteins are able to fold spontaneously and directly to their native state, some larger and multi-domain proteins, fold via multiple parallel pathways, at times engaging with competing basins of attraction, existing as an ensemble of misfolded structures known as non-native states. Proteins that become stuck in non-native states are prone to forming aggregates, due to exposed hydrophobes. Such aggregates have been implicated in various neurological diseases such as Parkinson's and Alzheimer's. Nature has evolved a protein quality control system which employs biological nanomachines, working to moderate off-pathway reactions, thereby modulating a balance between native and non-native proteins. There are two components to this system: protein degradation and protein folding assistance. While various components of this system engage in sundry tasks, I have focused on a unique system from each subsystem: p97, having unfolding activity and GroEL, having protein-folding assistance activity. p97, a ubiquitous hexameric, dual-ring ATPase found in eukaryotes, utilizes its high ATPase-activity D2 domains to drive dramatic allosteric conformational changes between distinct nucleotide-bound states. This results in paddling of highly-conserved loops found in the interior of the ring to pull proteins through its narrow central pore by repetitive engagement and release. Translocation of a substrate protein through this pore leads to unfolding, in preparation for proteolysis. I have used normal mode analysis to investigate how the collective motions of the p97 monomer, dimer and hexamer underlie p97's allosteric transitions. p97's open conformation shows common behavior in the two most-dominant collective motions in each system. Namely, a clockwise torsional motion suitable for directionality of substrate handling and an axial swing-like motion that transmits force to the substrate. Additionally, I looked at the directional correlation of the collective motions, which reveals a sequential nature of p97 and further suggests CW directionality in the dimer. Further, I performed a structural perturbation method, probing each residue’s energetic contribution. Coupling this to the aforementioned results, along with details of the collective motions of the systems, reveals a network of 23 residues which are important to the allosteric signaling network of p97. The chaperonins, providing protein-folding assistance via forced unfolding and sequestration of substrates, comprise two groups with distinct kinetics: the bacterial Group I utilize concerted allosteric motions, while the eukaryotic Group II utilizes sequential. I investigated protein folding coupled to the sequential allostery of a mutant bacterial GroEL using Langevin dynamics and a de novo candidate for folding assistance with a coarse-grained model. The sequential mutant shows dramatically reduced encapsulation ability. The confined substrate unfolding outcome is unchanged from wild-type. These data suggest encapsulation may not be a requirement for sequential allosteric protein folding assistance.
George Stan, Ph.D. (Committee Chair)
Thomas Beck, Ph.D. (Committee Member)
Pearl Tsang, Ph.D. (Committee Member)
98 p.

Recommended Citations

Citations

  • Smith, N. B. (2015). Computational Studies of Protein Folding Assistance and Conformational Pathways of Biological Nanomachines [Doctoral dissertation, University of Cincinnati]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1448037580

    APA Style (7th edition)

  • Smith, Nathan. Computational Studies of Protein Folding Assistance and Conformational Pathways of Biological Nanomachines. 2015. University of Cincinnati, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=ucin1448037580.

    MLA Style (8th edition)

  • Smith, Nathan. "Computational Studies of Protein Folding Assistance and Conformational Pathways of Biological Nanomachines." Doctoral dissertation, University of Cincinnati, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1448037580

    Chicago Manual of Style (17th edition)