Investigating strain dependency in the production of aromatic compounds in Saccharomyces cerevisiae.
Although Saccharomyces cerevisiae is the most highly domesticated yeast, strain dependency in biotechnological processes still remains as a common, yet poorly understood phenomenon. To investigate this, the entrance to the aromatic amino acid biosynthetic pathway was compared in four commonly used S. cerevisiae laboratory strains. The strains were engineered to accumulate shikimate by overexpressing a mutant version of the pentafunctional ARO1 enzyme with disrupted activity in the shikimate kinase subunit. Carbon tracing and13C metabolic flux analysis combined with quantitative PCR, revealed that precursor availability and shikimate production were dramatically different in the four equally engineered strains, which were found to be correlated with the strains' capacity to deal with protein overexpression burden. By implementing a strain-dependent approach, the genetic platform was reformulated, leading to an increase in yield and titer in all strains. The highest producing strain, INVSc1-SA3, produced 358 mg L-1of shikimate with a yield of 17.9 mg g-1glucose.These results underline the importance of strain selection in developing biological manufacturing processes, demonstrate the first case of high production of shikimate in yeast, and provide an appropriate platform for strain selection for future production of aromatic compounds. Biotechnol. Bioeng. 2016;113: 2676-2685. © 2016 Wiley Periodicals, Inc.
Department of Chemical and Biological Engineering, Iowa State University, Ames, Iowa.|NSF Engineering Research Center for Biorenewable Chemicals (CBiRC), Ames, Iowa.
Miguel
M
Suástegui
Department of Biological Systems Engineering, Virginia Polytechnic Institute and State University, Blacksburg, Virginia.
Weihua
W
Guo
Department of Biological Systems Engineering, Virginia Polytechnic Institute and State University, Blacksburg, Virginia. xueyang@vt.edu.
Xueyang
X
Feng
Department of Chemical and Biological Engineering, Iowa State University, Ames, Iowa. zyshao@iastate.edu.|NSF Engineering Research Center for Biorenewable Chemicals (CBiRC), Ames, Iowa. zyshao@iastate.edu.
Zengyi
Z
Shao
D003198:Computer Simulation; D015966:Gene Expression Regulation, Fungal / Q000235:genetics; D006841:Hydrocarbons, Aromatic / Q000302:isolation & purification / Q000378:metabolism; D060847:Metabolic Engineering / Q000379:methods*; D064688:Metabolic Flux Analysis / Q000379:methods*; D008954:Models, Biological; D012441:Saccharomyces cerevisiae / Q000145:classification* / Q000235:genetics* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D012765:Shikimic Acid / Q000302:isolation & purification / Q000378:metabolism*; D013045:Species Specificity
D006841:Hydrocarbons, Aromatic; D029701:Saccharomyces cerevisiae Proteins; D012765:Shikimic Acid
13C-metabolic flux analysis; pathway balancing; protein overexpression burden; shikimate pathway