ADVANCING WORKFLOWS FOR STRUCTURE DETERMINATION OF DISEASE-RELEVANT MACRO-MOLECULES
Restricted (Penn State Only)
- Author:
- Jonaid, G M
- Graduate Program:
- Bioinformatics and Genomics (PhD)
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- July 16, 2024
- Committee Members:
- Dajiang Liu, Outside Unit Member
Susan Hafenstein, Outside Field Member
David Koslicki, Program Head/Chair
David Koslicki, Chair & Dissertation Advisor
Santhosh Girirajan, Major Field Member - Keywords:
- Bioinformatics
data analysis - Abstract:
- Cryo-electron microscopy (cryo-EM) has revolutionized the field of structural biology by enabling the determination of high-resolution structures of macromolecules in their near-native states. This technology has opened up new avenues for understanding the molecular mechanisms of biological processes and has had a profound impact on drug discovery and development. In this work, we leveraged the power of cryo-EM to determine the high-resolution structure of the bacterial ATPase PilB, which plays a critical role in bacterial motility. Here, we report the cryo-EM structure of PilB, a bacterial ATPase essential for type IV pilus biogenesis and bacterial motility. The cryo-EM structure reveals the overall architecture of PilB and provides insights into the molecular mechanisms of ATP hydrolysis and conformational changes associated with PilB function. These findings provide a structural basis for understanding the mechanism of PilB function and offer potential targets for developing antimicrobial agents. In addition to cryo-EM, we attempted liquid-electron microscopy (liquid-EM) to visualize biological macromolecules in their native state. Our liquid-EM approach involves encapsulating biological molecules within a hermetically sealed chamber, safeguarding the samples from high vacuum conditions during imaging. While we attempted the visualization of adeno-associated virus (AAV) in liquid, the acquisition of data was hindered by beam damage and particle movement. Despite the current limitations of liquid-EM for structural biology, we present a pipeline encompassing sample preparation, data collection, and data processing that could prove valuable if future endeavors successfully address the challenges of beam damage and particle movement in liquid solution. Once these limitations are overcome, this technique holds the potential to unlock the visualization of previously inaccessible states of macromolecules in real-time manner.