Construction of a novel MK-4 biosynthetic pathway in Pichia pastoris through heterologous expression of HsUBIAD1

doi:10.1186/s12934-019-1215-9

	Construction of a novel MK-4 biosynthetic pathway in Pichia pastoris through heterologous expression of HsUBIAD1
	Sun,Xiaowen 1,2; Liu,Hui 2; Wang,Peng 2; wang,Li 2; Ni,Wenfeng 1,2; Yang,Qiang 1,2; Wang,Han 1,2; Tang,Hengfang 1,2; Zhao,Genhai 2; Zheng,Zhiming 2
刊名	Microbial Cell Factories
	2019-10-10
卷号	18
关键词	Pichia pastoris Aromatic prenyltransferase HsUBIAD1 Geranylgeranyl diphosphate synthase Menaquinone
ISSN号	1475-2859
DOI	10.1186/s12934-019-1215-9
通讯作者	Zhao,Genhai(zhgh327@ipp.ac.cn) ; Zheng,Zhiming(zhengzhiming2014@163.com)
英文摘要	AbstractBackgroundWith a variety of physiological and pharmacological functions, menaquinone is an essential prenylated product that can be endogenously converted from phylloquinone (VK1) or menadione (VK3) via the expression of Homo sapiens UBIAD1 (HsUBIAD1). The methylotrophic yeast, Pichia pastoris, is an attractive expression system that has been successfully applied to the efficient expression of heterologous proteins. However, the menaquinone biosynthetic pathway has not been discovered in P. pastoris.ResultsFirstly, we constructed a novel synthetic pathway in P. pastoris for the production of menaquinone-4 (MK-4) via heterologous expression of HsUBIAD1. Then, the glyceraldehyde-3-phosphate dehydrogenase constitutive promoter (PGAP) appeared to be mostsuitable for the expression of HsUBIAD1 for various reasons. By optimizing the expression conditions of HsUBIAD1, its yield increased by 4.37 times after incubation at pH 7.0 and 24?°C for 36?h, when compared with that under the initial conditions. We found HsUBIAD1 expressed in recombinant GGU-23 has the ability to catalyze the biosynthesis of MK-4 when using VK1 and VK3 as the isopentenyl acceptor. In addition, we constructed a ribosomal DNA (rDNA)-mediated multi-copy expression vector for the fusion expression of SaGGPPS and PpIDI, and the recombinant GGU-GrIG afforded higher MK-4 production, so that it was selected as the high-yield strain. Finally, the yield of MK-4 was maximized at 0.24?mg/g DCW by improving the GGPP supply when VK3 was the isopentenyl acceptor.ConclusionsIn this study, we constructed a novel synthetic pathway in P. pastoris for the biosynthesis of the high value-added prenylated product MK-4 through heterologous expression of HsUBIAD1 and strengthened accumulation of GGPP. This approach could be further developed and accomplished for the biosynthesis of other prenylated products, which has great significance for theoretical research and industrial application.
语种	英语
出版者	BioMed Central
WOS记录号	BMC:10.1186/S12934-019-1215-9
内容类型	期刊论文
源URL	[http://ir.hfcas.ac.cn:8080/handle/334002/42808]
专题	中国科学院合肥物质科学研究院
通讯作者	Zhao,Genhai; Zheng,Zhiming
作者单位	1. 2.
推荐引用方式 GB/T 7714	Sun,Xiaowen,Liu,Hui,Wang,Peng,et al. Construction of a novel MK-4 biosynthetic pathway in Pichia pastoris through heterologous expression of HsUBIAD1[J]. Microbial Cell Factories,2019,18.
APA	Sun,Xiaowen.,Liu,Hui.,Wang,Peng.,wang,Li.,Ni,Wenfeng.,...&Zheng,Zhiming.(2019).Construction of a novel MK-4 biosynthetic pathway in Pichia pastoris through heterologous expression of HsUBIAD1.Microbial Cell Factories,18.
MLA	Sun,Xiaowen,et al."Construction of a novel MK-4 biosynthetic pathway in Pichia pastoris through heterologous expression of HsUBIAD1".Microbial Cell Factories 18(2019).