Synthesis and release of the bacterial compatible solute 5-hydroxyectoine in Hansenula polymorpha

Abstract

Ectoine and 5-hydroxyectoine belong to the family of compatible solutes which are known to mainly contribute to the adaptation of the cell to osmotic stress by mediation of a constant turgor. In addition the cell's essential functions are maintained under stress conditions like high salinity, heat or aridity stress. Hansenula polymorpha was engineered to catalyze the transformation of monomeric substrates to 5-hydroxyectoine. For this purpose four genes encoding the enzymes of the 5-hydroxyectoine biosynthesis pathway of Halomonas elongata, EctA, EctB, EctC, and EctD, were inserted into the genome of H. polymorpha. Subsequently the syntheses of ectoine and 5-hydroxyectoine were analyzed and optimized. We showed that H. polymorpha is a suitable system for recombinant 5-hydroxyectoine synthesis in gram per liter scale (2.8 g L⁻¹ culture supernatant, 365 μmol/g dcw) in which almost 100% conversion of ectoine to 5-hydroxyectoine without necessity of high salinity were achieved.

Keywords: 5-Hydroxyectoine; Asd; Ask; Compatible solute; DABA; DABA acyltransferase; DABA-2-oxoglutarate transaminase; EctA; EctB; EctC; EctD; Ectoine; FMD; Halomonas elongata; Hansenula polymorpha; MOX; MeOH; Whole-cell biocatalysis; aspartate kinase; aspartate semialdehyde dehydrogenase; dcw; diaminobutyrate; dry cell weight; ectoine hydroxylase; ectoine synthase; formate dehydrogenase; methanol; methanol oxidase.