期刊
MICROBIAL CELL FACTORIES
卷 14, 期 -, 页码 -出版社
BMC
DOI: 10.1186/s12934-014-0187-z
关键词
Pichia pastoris; Recombinant protein production; Plant peroxidase; Horseradish peroxidase; Metabolic engineering; Cofactor; Heme; Heme biosynthesis; Apo-protein
资金
- NAWI Graz
- Austrian Science Fund FWF [P24861-B19, W901]
- European Union's Seventh Framework Programme (FP7) [266025]
- Austrian Science Fund (FWF) [P 24861] Funding Source: researchfish
- Austrian Science Fund (FWF) [P24861] Funding Source: Austrian Science Fund (FWF)
Background: Insufficient incorporation of heme is considered a central impeding cause in the recombinant production of active heme proteins. Currently, two approaches are commonly taken to overcome this bottleneck; metabolic engineering of the heme biosynthesis pathway in the host organism to enhance intracellular heme production, and supplementation of the growth medium with the desired cofactor or precursors thereof to allow saturation of recombinantly produced apo-forms of the target protein. In this study, we investigated the effect of both, pathway engineering and medium supplementation, to optimize the recombinant production of the heme protein horseradish peroxidase in the yeast Pichia pastoris. Results: In contrast to studies with other hosts, co-overexpression of genes of the endogenous heme biosynthesis pathway did not improve the recombinant production of active heme protein. However, medium supplementation with hemin proved to be an efficient strategy to increase the yield of active enzyme, whereas supplementation with the commonly used precursor 5-aminolevulinic acid did not affect target protein yield. Conclusions: The yield of active recombinant heme peroxidase from Pichia pastoris can be easily enhanced by supplementation of the cultivation medium with hemin. Thereby, secreted apo-species of the target protein are effectively saturated with cofactor, maximizing the yield of target enzyme activity.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据