Master of Science Amalie Skogvold will Friday March 6th, 2026, at 12:15 hold her Thesis Defense for the PhD degree in Science. The title of the thesis is:
« Enzymes in the biosynthesis- and degradation pathways of ectoine »
Microbial survival in high salt habitats depends on maintaining osmotic balance, matching cytoplasmic solute concentrations to external salinity to prevent water loss. One strategy is the synthesis or uptake of compatible solutes such as ectoine and 5-hydroxyectoine, which protect cells without disrupting core metabolism. This thesis characterizes enzymes that synthesize and degrade ectoine through structural and biochemical analyses. The main methods used were X-ray crystallography, complemented by SEC-MALS for oligomeric state, DSC and Prometheus for thermal stability, coupled activity assays, site directed mutagenesis, and molecular docking. High resolution structures were determined from both pathways. The degradation transaminase DoeD from Chromohalobacter salexigens was determined to be a fold type I PLP dependent transaminase, and a homodimer with PLP covalently bound to an active site lysine residue. Enzyme assays established pyruvate and alanine as the co-substrate pair, revising a common previous model. The transaminase DoeD had activity toward several ω-amino acids but also the α-amino acid homoserine. The biosynthetic transaminase EctB from cold adapted Marinobacter sp. CK1 was shown to be a homotetramer. Structures of the wild type and an inactive K264A variant, together with mutational analysis, demonstrated that tetramerization is required both for activity and to improve thermal stability, and also identified catalytic residues including Tyr14 and Arg295 that are critical for activity. Similar to DoeD, EctB had activity toward several ω-amino acids, but not homoserine. The structure of ectoine synthase EctC from C. salexigens was solved and showed a chloride ion near the active site and β-hydroxylation of an Asn40 involved in substrate positioning. Biochemical and thermal analysis revealed a stability-activity trade off in which activity peaks at basic pH while stability is higher near neutral pH, with further stabilization by salt and Fe2+. The findings of this work link quaternary structure to transaminase performance and provide a structural basis for engineering robust biocatalysts for improved ectoine production and green chiral amine synthesis.
1st Opponent: Dr. Andy-Mark Thunnissen, Groningen Biomolecular Sciences and Biotechnology Institute (GBB), University of Groningen, Groningen, The Netherlands
2nd Opponent: Professor Aurora Martinez, University of Bergen, The Department of Biomedicine, Bergen, Norway
Internal member and leader of the committee: Professor Peik Haugen, Department of chemistry, NT-Fak, UiT