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Biochemical and Biophysical Characterization of Huntingtin


Owens, Gwen Ellen (2016) Biochemical and Biophysical Characterization of Huntingtin. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/Z9TM7835.


Huntington’s disease (HD) is a fatal autosomal dominant neurodegenerative disease. HD has no cure, and patients pass away 10-20 years after the onset of symptoms. The causal mutation for HD is a trinucleotide repeat expansion in exon 1 of the huntingtin gene that leads to a polyglutamine (polyQ) repeat expansion in the N-terminal region of the huntingtin protein. Interestingly, there is a threshold of 37 polyQ repeats under which little or no disease exists; and above which, patients invariably show symptoms of HD. The huntingtin protein is a 350 kDa protein with unclear function. As the polyQ stretch expands, its propensity to aggregate increases with polyQ length. Models for polyQ toxicity include formation of aggregates that recruit and sequester essential cellular proteins, or altered function producing improper interactions between mutant huntingtin and other proteins. In both models, soluble expanded polyQ may be an intermediate state that can be targeted by potential therapeutics.

In the first study described herein, the conformation of soluble, expanded polyQ was determined to be linear and extended using equilibrium gel filtration and small-angle X-ray scattering. While attempts to purify and crystallize domains of the huntingtin protein were unsuccessful, the aggregation of huntingtin exon 1 was investigated using other biochemical techniques including dynamic light scattering, turbidity analysis, Congo red staining, and thioflavin T fluorescence. Chapter 4 describes crystallization experiments sent to the International Space Station and determination of the X-ray crystal structure of the anti-polyQ Fab MW1. In the final study, multimeric fibronectin type III (FN3) domain proteins were engineered to bind with high avidity to expanded polyQ tracts in mutant huntingtin exon 1. Surface plasmon resonance was used to observe binding of monomeric and multimeric FN3 proteins with huntingtin.

Item Type:Thesis (Dissertation (Ph.D.))
Subject Keywords:Huntingtin; Huntington's disease; Polyglutamine; X-ray crystallography
Degree Grantor:California Institute of Technology
Division:Chemistry and Chemical Engineering
Major Option:Biochemistry and Molecular Biophysics
Awards:Gerald S. Levey, M.D., Medical Scientist Scholarship
Thesis Availability:Public (worldwide access)
Research Advisor(s):
  • Bjorkman, Pamela J.
Thesis Committee:
  • Rees, Douglas C. (chair)
  • Phillips, Robert B.
  • Zinn, Kai George
  • Bjorkman, Pamela J.
Defense Date:29 October 2015
Funding AgencyGrant Number
Center for the Advancement of Science in SpaceUNSPECIFIED
W. R. Hearst Endowed ScholarshipUNSPECIFIED
National Institutes of Health5T32GM007616
National Institutes of Health5T32GM008042
Record Number:CaltechTHESIS:11042015-142900908
Persistent URL:
Related URLs:
URLURL TypeDescription adapted for Ch. 2
Owens, Gwen Ellen0000-0003-0793-1994
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:9270
Deposited By: Gwen Owens
Deposited On:16 May 2016 16:51
Last Modified:08 Nov 2023 00:16

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