Expression, purification and structural characterization of human lysine-ketoglutarate reductase/Saccharopine dehydrogenase (LKR/SDH)

• Grant number: 13/12271-9
• Support Opportunities: Scholarships abroad - Research Internship - Doctorate (Direct)
• Start date: September 02, 2013
• End date: March 01, 2014
• Field of knowledge: Biological Sciences - Biochemistry - Chemistry of Macromolecules
• Principal Investigator: Paulo Arruda
• Grantee: Izabella Agostinho Pena
• Supervisor: Wyatt Yue
• Host Institution: Centro de Biologia Molecular e Engenharia Genética (CBMEG). Universidade Estadual de Campinas (UNICAMP). Campinas , SP, Brazil
• Institution abroad: University of Oxford, England
• Associated to the scholarship: 12/00235-5 - Mechanisms of saccharopine pathway induction in human cells, BP.DD

Abstract

The Saccharopine pathway of lysine degradation has been studied for decades in plants and mammals as the main regulator of lysine levels. It has been demonstrated that its role goes beyond the catabolic component, connecting to cellular responses against oxidative and osmotic stresses. The first steps are performed by the bifunctional enzyme LKR-SDH (Lysine Ketoglutarate Reductase (LKR)/ Saccharopine Dehydrogenase (SDH)) and Aminoadipate Semialdehyde Dehydrogenase (ALDH7A1). In mammals there are no studies using genetic manipulation for the phenotypic study of LKR-SDH inactivation. It has been known that mutations in the LKR-SDH gene can lead to hyperlysinemia, an inborn error of metabolism that can have clinical manifestation. In the other side, mutations in the human ALDH7A1 gene cause pyridoxine-dependent and folic acid-responsive seizures, a severe condition generated by the accumulation of aminoadipate semialdehyde (AASA). One possible therapy against these seizures could be the reduction of LKR/SDH activity, which would reduce AASA production. In order to do this, the biochemical and structural properties of human LKR/SDH should be deeply deciphered. In this context and aligned with my PhD project, we propose to express, purify and determine the structure of human LKR/SDH enzyme in collaboration with the group Metabolic & Rare Diseases of Structural Genomics Consortium (SGC) at Oxford University, United Kingdom, one of the main centers for studies in human protein structures in the world. This research internship will be supervised by Dr. Wyatt Yue, who is responsible for the group, due to its extensive experience with structural studies of metabolic protein. (AU)