title: Intracellular expression of Vitreoscilla hemoglobin improves production of Yarrowia lipolytica lipase LIP2 in a recombinant Pichia pastoris.
abstract: The Yarrowia lipolytica lipase LIP2 (YlLIP2) gene lip2 and Vitreoscilla hemoglobin gene vgb were co-expressed in Pichia pastoris, both under the control of AOX1 promoter, in order to alleviate respiration limitation under conditions of high cell-density fermentation and enhance YlLIP2 production. The results showed that recombinant P. pastoris strains harboring the lip2 and vgb genes (VHb(+)) displayed higher biomass and YlLIP2 activity than control strains (VHb(-)). Compared with VHb(-) cells, the expression levels of YlLIP2 in VHb-expressing cells when oxygen was not a limiting factor were improved 31.5% in shake-flask culture and 22% in a 10-L fermentor. Under non-limiting dissolved oxygen (DO) conditions, the maximum YlLIP2 activity of VHb(+) in a 10-L fermentor reached 33,000 U/mL. Oxygen limitation had a more negative effect on YlLIP2 productivity in VHb(-) cells than in VHb(+) cells. The highest YlLIP2 activity of VHb(+) cells was approximately 1.84-fold higher than that of VHb(-) cells at lower DO levels. Moreover, the recombinant strain VHb(+) exhibited a higher specific oxygen uptake rate and achieved higher cell viability under oxygen limiting and non-limiting conditions compared with VHb(-) cells. Therefore, the above results suggest that intracellular expression of VHb in recombinant P. pastoris has the potential to improve cell growth and industrial enzyme production.
authors: Key Laboratory of Molecular Biophysics of the Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan 430074, China.
authors: Xiaofeng
authors: X
authors: Wang
authors: Yongchuan
authors: Y
authors: Sun
authors: Xuguang
authors: X
authors: Shen
authors: Feng
authors: F
authors: Ke
authors: Heyun
authors: H
authors: Zhao
authors: Yun
authors: Y
authors: Liu
authors: Li
authors: L
authors: Xu
authors: Yunjun
authors: Y
authors: Yan
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D019149:Bioreactors / Q000382:microbiology; D017931:DNA Primers / Q000235:genetics; D005285:Fermentation; D005656:Fungal Proteins / Q000096:biosynthesis* / Q000235:genetics*; D015870:Gene Expression; D005798:Genes, Bacterial; D005800:Genes, Fungal; D008049:Lipase / Q000096:biosynthesis* / Q000235:genetics*; D010843:Pichia / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D054793:Truncated Hemoglobins / Q000235:genetics* / Q000378:metabolism*; D020589:Vitreoscilla / Q000235:genetics* / Q000378:metabolism*; D025062:Yarrowia / Q000201:enzymology / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; D005656:Fungal Proteins; D011994:Recombinant Proteins; D054793:Truncated Hemoglobins; C082613:hemoglobin protein, Vitreoscilla; C408971:LIP2 protein, Yarrowia lipolytica; D008049:Lipase