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Expression, Purification and Structural Characterization of Zebrafish Mitochondrial Serine Hydroxymethyltransferase
Thesis

Expression, Purification and Structural Characterization of Zebrafish Mitochondrial Serine Hydroxymethyltransferase

Chen, Yi-Hung
Masters, 國立清華大學, 物理系
2012

Abstract

斑馬魚 絲胺酸羥基甲基轉移酵素 同步輻射圓二色圖譜儀 動態光散射粒徑分析儀 小角度 X 光散射實驗 Zebrafish Serine Hydroxymethyltransferase SRCD DLS SAXS
Serine hydroxymethyltransferase (SHMT), widely distributed in nature including Prokaryotic and Eukaryotic organisms, catalyzes several important biosynthesis reactions, such as in folate metabolism. Functions of SHMTs have been extensively studied in many living systems, and enzymic imbalance in serine and folate metabolism have been reported to cause diseases including cancer. SHMT can be target for designing and developing inhibitors which could be used as anticancer drugs. The zebrafish has risen to be a prominent animal model because it is easy to grow and breed and has comparable organs and tissues with mammals. Zebrafish Mitochondrial Serine Hydroxymethyltransferase(zmSHMT) has been expressed in E. coli and studied, but structural information remains unknown. In this thesis, Dynamic Light Scattering(DLS) serves as homogeneity indicator during purification, Synchrotron Radiation Circular Dichroism(SRCD) helps identifying secondary structures, and Small-Angel X-ray Scattering(SAXS) helps calculating hydrodynamic radius information and resolving low resolution structural information in solution. With above results, zmSHMT shows more $\beta$-sheet secondary structure and larger size than that of other existing structures of Eukaryotic species including human, rabbit and mouse, which also imply elongated tertiary structure that may lead to differences from mammals.

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