STRUCTURAL CHARACTERIZATION OF BIOLOGICAL ASSEMBLIES USING ADVANCED X-RAY SCATTERING TECHNIQUES
Open Access
- Author:
- Ye, Dan
- Graduate Program:
- Chemical Engineering
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- May 29, 2018
- Committee Members:
- Enrique D. Gomez, Dissertation Advisor/Co-Advisor
Enrique D. Gomez, Committee Chair/Co-Chair
Esther W. Gomez, Committee Member
Seong Han Kim, Committee Member
Susan Hafenstein, Outside Member - Keywords:
- X-ray
Scattering
Bovine serum alubmin
Plant cell wall
Aluminum oxide - Abstract:
- In this dissertation, we explored the potential of using advanced X-ray scattering techniques to examine the 3-D structure of subcomponents in biological assemblies. The majority of this dissertation focuses on resonant soft X-ray scattering (RSoXS). The goal is to extend soft X-rays to characterize biological assemblies. We conducted proof of concept experiments on bovine serum albumin (BSA) in solution. We demonstrated that, with 1000 times less scattering volume than traditional hard X-rays, RSoXS is able to provide the size and shape of BSA molecules. More importantly, we show that different components of BSA are highlighted when incident beam energies are tuned to highlight specific chemical moieties. We also examined the spacing of cellulose microfibrils in onion epidermal cell walls. Quantification of the spacing between cellulose microfibrils in primary cell walls is still challenging using microscopy or SAXS. To enhance contrast, calcium ions which are bind to the pectin surrounding the cellulose microfibrils were introduced to the system. We found that the average interfibril spacing between microfibrils is on average 20 nm in onion epidermal cell walls near Ca L-edge. We also designed and fabricated aluminum oxide thin films as the next generation sample substrates for N RSoXS. The Al2O3 windows offer higher X-ray transmittance and have no fluorescence background near N K-edge. Furthermore, the Al2O3 windows allow the 30 nm domain spacing from a block copolymer to be revealed near N K-edge which could not be observed using commercially available Si3N4 windows. Altogether, our findings indicate that RSoXS can be extended to characterize more complexed biological systems such as membrane proteins and viruses in solution. In addition, we investigated the cellulose crystal orientation in onion epidermal cell walls using grazing incident wide angle x-ray scattering (GIWAXS). In constrast to the predominant assumption that the cellulose crystallites are twisted helically along cellulose microfibrils, the GIWAXS result indicates the cellulose crystals have strong out-of-plane crystalline order but poor in-plane crystalline order. Our results imply that the preferred orientation of cellulose crystals could be related to hydrophobic and hydrophilic interactions in primary cell walls.