The formation of elaborate molecular structures is a hallmark of biological systems, resulting in enzyme networks, regulatory complexes, membrane signaling and packaging systems, scaffolding networks, and . The instructions for assembly are encoded within the molecules themselves. Understanding how to read these instructions is a major goal in modern biology and in modern medicine, where understanding may lead to rational manipulation of disease mechanisms. One relatively simple yet no less spectacular self-assembly process is the protein folding problem. Understanding the rules, rates, and mechanisms of protein folding would help interpret all aspects of biology, including disease states relating to protein misfolding and aggregation. One of the central features of protein folding is cooperativity. Single-domain proteins fold in all-or-none reactions. This remarkable coupling phenomenon is likely to be important for avoiding misfolded/partly folded states in biology, but it complicates experimental studies of the folding process, masking intermediates, processes, and routes along the way to the native state. The proposed research uses a series of linear symmetric repeat proteins that display all the features of globular proteins, but greatly reduce the complexity of the problem, and allow nearest-neighbor models to be applied to quantify the energetic coupling that underlies cooperativity. This internal symmetry also permits detailed kinetic mechanisms to be tested and parameterized, including parallel pathway nucleation and propagation steps. These modes of analysis will be applied alpha helix and beta sheet containing repeats to measure cooperativity and nucleation kinetic processes and determine their structural origins. Using a recently identified length-variable set of repeats, the size of the repeats and their interfaces with their neighbors will be correlated to cooperativity terms. Further, we will delineate the sequence features that modulate length in this group of proteins, and explore the structural and energetic features of """"""""high-ID repeat protein sequences and their relation to consensus sequences.

Public Health Relevance

The proposed research will determine how parts of proteins come together to form stable structures with biological activities. We will achieve this by using a novel class of proteins wher we can snap units together like Legos (children's building blocks) and compare the structures, stabilities, and speeds with which they fold up. These results will help to explain how proteins avoid forming toxic, misfolded aggregates, and what has gone wrong when they do.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Special Emphasis Panel (ZRG1)
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Smith, Ward
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Johns Hopkins University
Schools of Arts and Sciences
United States
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Fossat, Martin J; Dao, Thuy P; Jenkins, Kelly et al. (2016) High-Resolution Mapping of a Repeat Protein Folding Free Energy Landscape. Biophys J 111:2368-2376
Geiger-Schuller, Kathryn; Barrick, Doug (2016) Broken TALEs: Transcription Activator-like Effectors Populate Partly Folded States. Biophys J 111:2395-2403
Marold, Jacob D; Kavran, Jennifer M; Bowman, Gregory D et al. (2015) A Naturally Occurring Repeat Protein with High Internal Sequence Identity Defines a New Class of TPR-like Proteins. Structure 23:2055-65
Dao, Thuy Phuong; Majumdar, Ananya; Barrick, Doug (2015) Highly polarized C-terminal transition state of the leucine-rich repeat domain of PP32 is governed by local stability. Proc Natl Acad Sci U S A 112:E2298-306
Preimesberger, Matthew R; Majumdar, Ananya; Aksel, Tural et al. (2015) Direct NMR detection of bifurcated hydrogen bonding in the α-helix N-caps of ankyrin repeat proteins. J Am Chem Soc 137:1008-11
Dao, Thuy P; Majumdar, Ananya; Barrick, Doug (2014) Capping motifs stabilize the leucine-rich repeat protein PP32 and rigidify adjacent repeats. Protein Sci 23:801-11
Aksel, Tural; Barrick, Doug (2014) Direct observation of parallel folding pathways revealed using a symmetric repeat protein system. Biophys J 107:220-32
Cunha, Eva S; Hatem, Christine L; Barrick, Doug (2013) Insertion of endocellulase catalytic domains into thermostable consensus ankyrin scaffolds: effects on stability and cellulolytic activity. Appl Environ Microbiol 79:6684-96
Aksel, Tural; Majumdar, Ananya; Barrick, Doug (2011) The contribution of entropy, enthalpy, and hydrophobic desolvation to cooperativity in repeat-protein folding. Structure 19:349-60
Vieux, Ellen F; Barrick, Doug (2011) Deletion of internal structured repeats increases the stability of a leucine-rich repeat protein, YopM. Biophys Chem 159:152-61

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