Fig. 1: Engineering a THIQ-producing yeast.
From: A yeast platform for high-level synthesis of tetrahydroisoquinoline alkaloids
a (S)-Norcoclaurine (3) synthetic pathway in engineered yeast. The native yeast Ehrlich pathway (green) supplies 4-HPAA (1) from l-tyrosine, while a heterologous pathway (blue) generates dopamine (2), also from l-tyrosine. CYP76AD5 is a more active plant tyrosine hydroxylase compared to CYP76AD1* (CYP76AD1W13L F309L), which is an engineered variant of CYP76AD1. NCS catalyzes an enantioselective Pictet-Spengler condensation of 4-HPAA and dopamine, yielding (S)-norcoclaurine (pink). Native shikimate and l-tyrosine biosynthetic pathways are shown in orange. b (S)-Norcoclaurine (3) titer in culture supernatants of successive engineered strains. CjNCS stereo-specifically yields (S)-norcoclaurine64, while the stereo-specificity of NdNCS has not been reported. Error bars represent the mean ± s.d. of n = 3 independent biological samples. All strains exhibited a significant increase (P < 0.05) in (S)-norcoclaurine titer relative to the respective parental strain, with the exception of strain LP442, which exhibited a significant decrease (P < 0.05) in titer relative to strain LP412. Statistical differences between control and derivative strains were tested using two-tailed Student’s t-test. Refer to Supplementary Data 3 for genotypes of the full 23-strain (S)-norcoclaurine lineage. l-DOPA l-3,4-dihydroxyphenylalanine, DODC DOPA decarboxylase, E4P erythrose-4-phosphate, 4-HPAA 4-hydroxyphenylacetaldehyde, 4-HPAC 4-hydroxyphenylacetic acid, 4-HPP 4-hydroxyphenylpyruvate, NCS norcoclaurine synthase, PEP phosphoenolpyruvate, l-Phe l-phenylalanine, l-Trp l-tryptophan, l-Tyr l-tyrosine. Source data underlying Fig. 1b are provided in a Source Data file.
