Methods and systems to secrete lignin-modifying enzymes and uses thereof
View Patent ↗Embodiments of the invention are generally directed to lignin-modifying enzymes and systems and methods of their manufacture. In many embodiments, yeast strains, including S. cerevisiae , are used to produce and secrete lignin-modifying enzymes Further embodiments are directed to methods to screening peroxidase-producing yeast strains, including S. cerevisiae . Additional embodiments are directed to an expression vector or cassette encoding for a protein of interest and one or more proteins to allow for surface to display of the protein of interest.
1 . A transgenic yeast organism comprising:
a knockout of a first native gene and a knockout of a second native gene, wherein the first native gene is cyt2 encoding cytochrome c heme lyase and the second native gene is pmt2 encoding protein O-mannosyltransferase 2; and
an expression cassette comprising a nucleotide sequence encoding for a lignin-modifying enzyme.
2 . The transgenic yeast organism of claim 1 , wherein the lignin-modifying enzyme is a heme peroxidase.
3 . The transgenic yeast organism of claim 1 , wherein the nucleotide sequence encoding the lignin-modifying enzyme is one of SEQ ID NOs: 1-78.
4 . The transgenic yeast organism of claim 1 , wherein the expression cassette further comprises a nucleotide sequence encoding a scaffold protein and a surface display protein, wherein the surface display protein is linked to the lignin-modifying enzyme, such that a hybrid peptide is produced with the lignin-modifying enzyme and the surface display protein.
5 . The transgenic yeast organism of claim 1 , wherein the expression cassette further comprises an inducible promoter operatively linked to the nucleotide sequence encoding the lignin-modifying enzyme.