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Mechanisms of activation and inhibition of Arp2/3 complex / Austin J. Zimmet.
- Format:
- Book
- Thesis/Dissertation
- Author/Creator:
- Zimmet, Austin J., author.
- Language:
- English
- Subjects (All):
- Biochemistry.
- Cellular biology.
- Molecular biology.
- Biochemistry and Molecular Biophysics--Penn dissertations.
- Penn dissertations--Biochemistry and Molecular Biophysics.
- Local Subjects:
- Biochemistry.
- Cellular biology.
- Molecular biology.
- Biochemistry and Molecular Biophysics--Penn dissertations.
- Penn dissertations--Biochemistry and Molecular Biophysics.
- Genre:
- Academic theses.
- Physical Description:
- 1 online resource (189 pages)
- Contained In:
- Dissertations Abstracts International 82-07B.
- Place of Publication:
- [Philadelphia, Pennsylvania] : University of Pennsylvania ; Ann Arbor : ProQuest Dissertations & Theses, 2020.
- Language Note:
- English
- System Details:
- Mode of access: World Wide Web.
- text file
- Summary:
- Arp2/3 complex nucleates branched actin networks that drive cell motility. It consists of seven proteins, including actin-related proteins (Arps) 2 and 3. In isolation Arp2/3 complex is in an inactive, splayed conformation. Two nucleation-promoting factors (NPFs) bind Arp2/3 complex during activation, but the order, specific interactions and contribution of each NPF to activation are unresolved. Here, we report the cryo-EM structure of recombinantly-expressed human Arp2/3 complex with two WASP-family NPFs bound and address the mechanism of activation. A crosslinking assay that captures the transition of the Arps into the activated filament-like conformation shows that actin binding to NPFs favors this transition. Actin-NPF binding to Arp2 precedes binding to Arp3 and is sufficient to promote the filament-like conformation, but not activation. Structure-guided mutagenesis of the NPF-binding sites reveals their distinct roles in activation and shows that, contrary to budding yeast Arp2/3 complex, NPF-mediated delivery of actin at the barbed end of both Arps is required for activation of human Arp2/3 complex.In addition to the complex being activated by NPFs, the complex is negatively regulated by a homologous protein, Arpin. Here, we also report the cryo-EM structure of bovine Arp2/3 complex bound to human Arpin. We've shown that despite its sequence similarity to the NPFs, Arpin specifically binds to a single binding site on Arp3. We have also shown that unlike GMF, Arpin acts on both ADP and ATP Arp2/3 complex. Here we have identified a unique mechanism of inhibition on Arp2/3 complex by Arpin.
- Notes:
- Source: Dissertations Abstracts International, Volume: 82-07, Section: B.
- Advisors: Dominguez, Roberto; Committee members: E. Ostap; Kenji Murakami; Tatyana Svitkina; Shae Padrick.
- Department: Biochemistry and Molecular Biophysics.
- Ph.D. University of Pennsylvania 2020.
- Local Notes:
- School code: 0175
- ISBN:
- 9798557065047
- Access Restriction:
- Restricted for use by site license.
- This item is not available from ProQuest Dissertations & Theses.
- This item must not be sold to any third party vendors.
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