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Obtainment, selection and characterization of a mutant strain of Kluyveromyces marxianus that displays improved production of 2‐phenylethanol and enhanced DAHP synthase activity
Author(s) -
Lima L.A.,
Ventorim R.Z.,
Bianchini I.A.,
Queiroz M.V.,
Fietto L.G.,
Silveira W.B.
Publication year - 2021
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1111/jam.14793
Subject(s) - mutant , kluyveromyces marxianus , atp synthase , biochemistry , strain (injury) , biology , kluyveromyces , yeast , enzyme , biosynthesis , gene , saccharomyces cerevisiae , anatomy
Aims Yeasts produce 2‐phenylethanol (2‐PE) from sugars via de novo synthesis; however, its synthesis is limited due to feedback inhibition on the isofunctional 3‐deoxy‐ d ‐arabino‐heptulosonate‐7‐phosphate (DAHP) synthases (Aro3p and Aro4p). This work aimed to select Kluyveromyces marxianus mutant strains with improved capacity to produce 2‐PE from sugars. Methods and Results Kluyveromyces marxianus CCT 7735 mutant strains were selected from UV irradiation coupled with screening of p‐ fluoro‐ dl ‐phenylalanine (PFP) tolerant strains on culture medium without l ‐Phe addition. Most of them produced 2‐PE titres higher than the parental strain and the Km_PFP41 mutant strain stood out for displaying the highest 2‐PE specific production rate. Moreover it showed higher activity of DAHP synthase than the parental strain. We sequenced both ARO3 and ARO4 genes of Km_PFP41 mutant and identified mutations in ARO4 which caused changes in both size and conformation of the Aro4p. These changes seem to be associated with the enhanced activity of DAHP synthase and improved production of 2‐PE exhibited by that mutant strain. Conclusions The Km_PFP41 mutant strain presented improved 2‐PE production via de novo synthesis and enhanced DAHP synthase activity. Significance and Impact of the Study The mutant strain obtained in this work may be exploited as a yeast cell factory for high‐level synthesis of 2‐PE.