title: Metabolic Engineering of Probiotic Saccharomyces boulardii.
abstract: Saccharomyces boulardiiis a probiotic yeast that has been used for promoting gut health as well as preventing diarrheal diseases. This yeast not only exhibits beneficial phenotypes for gut health but also can stay longer in the gut than Saccharomyces cerevisiae Therefore, S. boulardiiis an attractive host for metabolic engineering to produce biomolecules of interest in the gut. However, the lack of auxotrophic strains with defined genetic backgrounds has hampered the use of this strain for metabolic engineering. Here, we report the development of well-defined auxotrophic mutants (leu2,ura3,his3, and trp1) through clustered regularly interspaced short palindromic repeat (CRISPR)-Cas9-based genome editing. The resulting auxotrophic mutants can be used as a host for introducing various genetic perturbations, such as overexpression or deletion of a target gene, using existing genetic tools forS. cerevisiae We demonstrated the overexpression of a heterologous gene (lacZ), the correct localization of a target protein (red fluorescent protein) into mitochondria by using a protein localization signal, and the introduction of a heterologous metabolic pathway (xylose-assimilating pathway) in the genome ofS. boulardii We further demonstrated that human lysozyme, which is beneficial for human gut health, could be secreted by S. boulardii Our results suggest that more sophisticated genetic perturbations to improveS. boulardii can be performed without using a drug resistance marker, which is a prerequisite for in vivo applications using engineeredS. boulardii.
authors: Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
authors: Jing-Jing
authors: JJ
authors: Liu
authors: Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.|Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
authors: In Iok
authors: II
authors: Kong
authors: Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
authors: Guo-Chang
authors: GC
authors: Zhang
authors: Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
authors: Lahiru N
authors: LN
authors: Jayakody
authors: Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.|Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
authors: Heejin
authors: H
authors: Kim
authors: Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.|School of Environmental Science and Engineering, Shandong University, Jinan, China.
authors: Peng-Fei
authors: PF
authors: Xia
authors: Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.|Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
authors: Suryang
authors: S
authors: Kwak
authors: Bioenergy and Biochemical Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, South Korea.
authors: Bong Hyun
authors: BH
authors: Sung
authors: Bioenergy and Biochemical Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, South Korea.
authors: Jung-Hoon
authors: JH
authors: Sohn
authors: Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
authors: Hanna E
authors: HE
authors: Walukiewicz
authors: Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
authors: Christopher V
authors: CV
authors: Rao
authors: Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA ysjin@illinois.edu.|Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
authors: Yong-Su
authors: YS
authors: Jin
mesh_terms: D015870:Gene Expression; D005827:Genetics, Microbial; D060847:Metabolic Engineering / Q000379:methods*; D008967:Molecular Biology / Q000379:methods; D019936:Probiotics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D012440:Saccharomyces / Q000235:genetics* / Q000254:growth & development
chemical_list: D011994:Recombinant Proteins