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Efficient production of S ‐adenosyl‐ l ‐methionine from dl ‐methionine in metabolic engineered Saccharomyces cerevisiae
Author(s) -
Liu Wei,
Tang Dandan,
Shi Rui,
Lian Jiazhang,
Huang Lei,
Cai Jin,
Xu Zhinan
Publication year - 2019
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.27157
Subject(s) - methionine , biochemistry , fermentation , saccharomyces cerevisiae , d amino acid oxidase , chemistry , amino acid , biology , yeast , enzyme , oxidase test
S ‐Adenosyl‐ l ‐methionine (SAM) is an important small molecule compound widely used in treating various diseases. Although l ‐methionine is generally used, the low‐cost dl ‐methionine is more suitable as the substrate for industrial production of SAM. However, d ‐methionine is inefficient for SAM formation due to the substrate‐specificity of SAM synthetase. In order to increase the utilization efficiency of dl ‐methionine, intracellular conversion of d ‐methionine to l ‐methionine was investigated in the type strain Saccharomyces cerevisiae BY4741 and an industrial strain S. cerevisiae HDL. Firstly, via disruption of HPA3 encoding d ‐amino acid‐ N ‐acetyltransferase, d ‐methionine was accumulated in vivo and no N ‐acetyl‐ d ‐methionine production was observed. Further, codon‐optimized d ‐amino acid oxidase ( DAAO ) gene from Trigonopsis variabilis (Genbank MK280686) and l ‐phenylalanine dehydrogenase gene ( l ‐PheDH ) from Rhodococcus jostii (Genbank MK280687) were introduced to convert d ‐methionine to l ‐methionine, SAM concentration and content was increased by 110% and 72.1% in BY4741 (plasmid borne) and increased by 38.2% and 34.1% in HDL (genome integrated), by feeding 0.5 g/L d ‐methionine. Using the recently developed CRISPR tools, the DAAO and l ‐PheDH expression cassettes were integrated into the HPA3 and SAH1 loci while SAM2 expression was integrated into the SPE2 and GLC3 loci of HDL, and the resultant strain HDL‐R2 accumulated 289% and 192% more SAM concentration and content, respectively, by feeding 0.5 g/L dl ‐methionine. Further, in a 10 L fed‐batch fermentation process, 10.3 g/L SAM were accumulated with the SAM content of 242 mg/g dry cell weight by feeding 16 g/L dl ‐methionine. The strategies used here provided a promising approach to enhance SAM production using low‐cost dl ‐methionine.