A synergistic effect on the production of S-adenosyl-L-methionine in Pichia pastoris by knocking in of S-adenosyl-L-methionine synthase and knocking out of cystathionine-beta synthase.
Two strategies of metabolic engineering have been used, individually or combined, to alter the metabolic flux to improve the production of S-adenosyl-L-methionine (SAM) in Pichia pastoris. One is over expressing SAM synthase by knock-in technique, the other is the disruption of cystathionine-beta synthase (CBS) by knock-out technique. Strain Gsam with ectopic SAM synthase gene produced 20 times of SAM comparing to the starter strain GS115. Disruption of CBS in GS115 only doubled its SAM production. However, disruption of CBS in Gsam results in a robust increase of SAM production, more than 56 times of the strain GS115. Thus, we report for the first time a synergistic effect on the production of SAM in yeast by the combination of knock-in and knock-out techniques. Furthermore, we optimize the cultural conditions for the genetically modified strain Gsam-cbs to produce SAM. The maximum yield of SAM reaches 3.6g/L in shake flask and 13.5g/L in a 5L fermentor, indicating that it could be used for industrial fermentation to produce large scale of SAM.
He
Junyun
J
National Key Laboratory of Protein Engineering and Plant Gene Engineering, LSC, Peking University, Beijing 100871, PR China.
Deng
Juanjuan
J
Zheng
Yinghua
Y
Gu
Jun
J
D019149:Bioreactors / Q000382:microbiology; D001709:Biotechnology / Q000379:methods; D003541:Cystathionine beta-Synthase / Q000235:genetics*; D005285:Fermentation; D005818:Genetic Engineering / Q000379:methods*; D008716:Methionine Adenosyltransferase / Q000235:genetics / Q000378:metabolism*; D010843:Pichia / Q000235:genetics*; D012436:S-Adenosylmethionine / Q000096:biosynthesis*
D012436:S-Adenosylmethionine; D008716:Methionine Adenosyltransferase; D003541:Cystathionine beta-Synthase