Cross Kingdom Glyco Protein Engineering

Publikation: Bog/antologi/afhandling/rapportPh.d.-afhandlingForskning

Svenning Rune Möller

This thesis covers our latest basic glycogene discovery efforts within plant O-glycosylation combined with our ongoing efforts to translate our findings into operational glycoengineering of plants which is only at the very beginning at this point in time. Pivotal to any cross species glycoengineering is detailed knowledge of the glycogenes and resulting structures involved which is provided in the two introductory chapters, where Chapter 1 ‘Plant and Mammalian Cell Protein N- and O-glycosylation – Recent Developments and Future Prospects’ provides a descriptive overview of N- and O-glycosylation structures and enzyme machineries primarily in mammalian cells and plants. The second part of the review, Chapter 2, ‘Trans kingdom glyco engineering - the biotechnological and pharmacological potential’, focuses on past and current efforts of controlling the two main types of protein glycosylation including the applicative potential hereof. Our extensin plant O-glycosylation machinery gene discovery efforts is described in Chapter 3 ‘Identification and evolution of a plant cell wall specific glycoprotein glycosyltransferase, ExAD’ and Chapter 4 ‘Complex regulation of Prolyl-4-hydroxylases impacts root hair expansion’, where the Arabidopsis arabinosyltransferase, AtExAD, has been shown to arabinosylate the plant cell wall glycoproteins, the extensins, and where the prolyl-4-hydroxylases, AtP4H2, -5, & -13, are functionally linked to hydroxylation of consecutive prolines in extensin repeat motifs, which is a prerequisite for the subsequent hydroxy proline arabinosylations. In Chapter 5, ‘Glycoproteomic analysis of seven major allergenic proteins reveals novel post-translational modifications ‘, we have, as part of a cross-disciplinary effort, participated in an in depth glycoprofiling analysis of seven major plant pollen allergens resulting the unraveling of an unprecedented and potentially adverse plant N- and O-linked glycosylation complexity, which may be involved in the allergnicity of the allergens. Chapter 6, ‘FACS medidated mutation enrichment of Nicotiana benthamiana protoplasts expressing agrobacterium delivered CRISPR/Cas9’, describes the use of viral replicons to deliver the CRISPR/Cas9 components to leaves of Nicotiana benthamiana, which we have optimized by fusing a Gfp marker to the Cas9 protein combined with FACS mediated cell sorting of Cas9-Gfp expressing protoplast cells.
ForlagDepartment of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen
StatusUdgivet - 2017

ID: 181414412