Co-ordinate expression of glycine betaine synthesis genes linked by the FMDV 2A region in a single open reading frame in Pichia pastoris.
The genes encoding the two enzymes choline monooxygenase (CMO) and betaine aldehyde dehydrogenase (BADH) of glycine betaine synthesis in Suaeda salsa were cloned and fused with the 2A region of foot-and-mouth disease virus in a single open reading frame. The fused genes were placed under the control of the alcohol oxidase (AOX1) promoter in pPIC3B and transformed into P. pastoris GS115. The expression of the fused genes in P. pastoris and the ability of recombinant yeasts to tolerate environmental stresses were studied. The results showed that induced with 0.5% methanol for 96 h, the maximal activities of CMO and BADH in the tested recombinant yeasts were 45- and 44-fold higher than those in the control yeast transformed empty vector only, respectively; the content of glycine betaine in the recombinant yeasts was 28- to 35-fold higher than that in the control. The fused genes linked by 2A region of foot-and-mouth disease virus were expressed in P. pastoris successfully and the polyprotein was 'cleaved' to each functional protein. The yeasts transformed the fused genes, which were more resistant to salt, methanol, and high temperature stresses than the control as result of glycine betaine synthesis genes introduced.
Wang
Sanhong
S
Laboratory of Fruit Tree Molecular Breeding, Horticulture Department, Nanjing Agricultural University, 6 Tongwei Rd, Nanjing, 210095, People's Republic of China.
Yao
Quanhong
Q
Tao
Jianmin
J
Qiao
Yushan
Y
Zhang
Zhen
Z
D000429:Alcohol Oxidoreductases / Q000235:genetics; D001622:Betaine / Q000378:metabolism*; D019663:Chenopodiaceae / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D029544:Foot-and-Mouth Disease Virus / Q000235:genetics / Q000378:metabolism; D016366:Open Reading Frames; D010105:Oxygenases / Q000096:biosynthesis* / Q000378:metabolism; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014764:Viral Proteins; C057421:virus protein 2A; D001622:Betaine; C105448:choline monooxygenase; D000429:Alcohol Oxidoreductases; C019643:alcohol oxidase; D010105:Oxygenases