Serial Femtosecond Crystallography of Proteins in Proteins and Cancer

• Other Authors: Ketawala, Gihan Kaushyal (Author)
• Format: Dissertation
• Language: English
• Published: 2020
• Subjects: Biochemistry; Mixed Lineage Leukemia; Photosynthesis; Photosystem II; Serial Femtosecond Crystallography; Taspase1; Time Resolved Crystallography
• Online Access: http://hdl.handle.net/2286/R.I.57193

abstract: This thesis focuses on serial crystallography studies with X-ray free electron lasers (XFEL) with a special emphasis on data analysis to investigate important processes in bioenergy conversion and medicinal applications. First, the work on photosynthesis focuses on time-resolved femtosecond crystallography studies of Photosystem II (PSII). The structural-dynamic studies of the water splitting reaction centering on PSII is a current hot topic of interest in the field, the goal of which is to capture snapshots of the structural changes during the Kok cycle. This thesis presents results from time-resolved serial femtosecond (fs) crystallography experiments (TR-SFX) where data sets are collected at room temperature from a stream of crystals that intersect with the ultrashort femtosecond X-ray pulses at an XFEL with the goal to obtain structural information from the transient state (S4) state of the cycle where the O=O bond is formed, and oxygen is released. The most current techniques available in SFX/TR-SFX to handle hundreds of millions of raw diffraction patterns are discussed, including selection of the best diffraction patterns, allowing for their indexing and further data processing. The results include two 4.0 Å resolution structures of the ground S1 state and triple excited S4 transient state. Second, this thesis reports on the first international XFEL user experiments in South Korea at the Pohang Accelerator Laboratory (PAL-XFEL). The usability of this new XFEL in a proof-of-principle experiment for the study of microcrystals of human taspase1 (an important cancer target) by SFX has been tested. The descriptions of experiments and discussions of specific data evaluation challenges of this project in light of the taspase1 crystals’ high anisotropy, which limited the resolution to 4.5 Å, are included in this report In summary, this thesis examines current techniques that are available in the SFX/TR-SFX domain to study crystal structures from microcrystals damage-free, with the future potential of making movies of biological processes. === Dissertation/Thesis === Masters Thesis Chemistry 2020