Evaluation of Brachypodium distachyon L-Tyrosine Decarboxylase Using L-Tyrosine Over-Producing Saccharomyces cerevisiae

Abstract

To demonstrate that herbaceous biomass is a versatile gene resource, we focused on the model plant Brachypodium distachyon, and screened the B. distachyon for homologs of tyrosine decarboxylase (TDC), which is involved in the modification of aromatic compounds. A total of 5 candidate genes were identified in cDNA libraries of B. distachyon and were introduced into Saccharomyces cerevisiae to evaluate TDC expression and tyramine production. It is suggested that two TDCs encoded in the transcripts Bradi2g51120.1 and Bradi2g51170.1 have L-tyrosine decarboxylation activity. Bradi2g51170.1 was introduced into the L-tyrosine over-producing strain of S. cerevisiae that was constructed by the introduction of mutant genes that promote deregulated feedback inhibition. The amount of tyramine produced by the resulting transformant was 6.6-fold higher (approximately 200 mg/L) than the control strain, indicating that B. distachyon TDC effectively converts L-tyrosine to tyramine. Our results suggest that B. distachyon possesses enzymes that are capable of modifying aromatic residues, and that S. cerevisiae is a suitable host for the production of L-tyrosine derivatives.

Structural basis for divergent and convergent evolution of catalytic machineries in plant aromatic amino acid decarboxylase proteins

“…TDC-clade enzymes from O. sativa (13), Camptotheca acuminate (14), C. annuum (12), Ophiorrhiza pumila (15), and C. roseus (16) have been shown to exhibit strict indolic substrate preference. On the contrary, previously characterized TyDC-clade enzymes Petroselinum crispum 4HPAAS (17), Rosa hybrid PAAS (18), Petunia hybrida PAAS (18), Rhodiola rosea 4HPAAS (10), Thalictrum flavum TyDC (17), Brachypodium distachyon TyDC (19) and P. somniferum TyDC (8) all exhibit phenolic substrate preference. It was noted that the indolic versus phenolic substrate preference of the TDC and TyDC clades are closely correlated with a predicted active-site-lining residue, which is conserved as a glycine within TDC clade but substituted to a serine or threonine within the TyDC clade (20).…”

Biosensor and machine learning-aided engineering of an amaryllidaceae enzyme

“…First, the central precursors 3,4-dihydroxybenzoate and tyramine must be overproduced in a base strain, likely Saccharomyces cerevisiae due to its proven ability to functionally express multiple plant-derived cytochrome proteins 44 . Biosynthetic pathways for the production of 3,4dihydroxybenzoate and tyramine have already been engineered into S. cerevisiae 45,46 , with high-throughput screening methods yielding improved tyrosine precursor yields 47 . Second, the early pathway enzymes for the production of norbelladine, norcraugsodine reductase, and norbelladine synthase must be functionally expressed in a microbial strain.…”

Synthetic microbial sensing and biosynthesis of amaryllidaceae alkaloids

“…First, the central precursors 3,4-dihydroxybenzoate and tyramine must be overproduced in a base strain, likely Saccharomyces cerevisiae due to its proven ability to functionally express multiple plant-derived cytochrome proteins 34 . Biosynthetic pathways for the production of 3,4-dihydroxybenzoate and tyramine have already been engineered into S. cerevisiae 35,36 , with high-throughput screening methods yielding improved tyrosine precursor yields 37 . Second, the early pathway enzymes for the production of norbelladine, norcraugsodine reductase, and norbelladine synthase must be functionally expressed in a microbial strain.…”