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Tailored nanodisc immobilization for one-step purification and reconstitution of cytochrome P450: A tool for membrane proteins' hard cases
Zhao, Lan1; Tao, Jiaoli1,2; Huang, Yongdong1; Zhu, Kai1; Du, Yuxiang1,2; Hao, Dongxia1; Liu, Hongying2; Zhang, Rongyue3; Ma, Guanghui1,4
2021-08-06
Source PublicationJOURNAL OF SEPARATION SCIENCE
ISSN1615-9306
Pages12
AbstractA novel nanodisc-based immobilization method was developed for high-efficient purification and reconstitution of cytochrome P450 in one step. Using membrane scaffold protein containing a histidine tag, charged-nanodiscs were prepared in the form of self-assembly of lipid-protein nanoparticles. Their properties including the particle diameter and its distribution and Zeta potential were controlled well by adjusting molar ratios of phospholipids to membrane scaffold protein. At an optimum lipid-to-membrane scaffold protein molar ratio of 60:1, uniformly regular-shaped and discoidal nanodiscs with an average particle diameter of 10 nm and Zeta potential of -19 mV were obtained. They can be well fractionated by size exclusion chromatography. Charged-nanodiscs were successfully immobilized onto Ni-chelating microspheres via histidine tags with a density of 6.6 mg membrane scaffold protein/mL gel. After being packed in a column, chromatography studies demonstrated that this nanodisc-immobilized chromatographic medium had a specific binding to cytochrome P450 in rat liver microsome. Nanodiscs containing cytochrome P450 can be furthermore eluted from the column with a diameter of about 87.0 nm and height of about 8.0 nm, respectively. The purity of cytochrome P450 after purification increased 25 folds strikingly. This nanodisc-immobilized chromatography method is promising for the one-step purification and reconstitution of membrane protein.
Keywordcytochrome P450 membrane protein purification nanodiscs nanodisc-immobilized chromatography reconstitution
DOI10.1002/jssc.202100284
Language英语
WOS KeywordKINETIC-ANALYSIS ; BINDING ; PERFORMANCE ; CHROMATOGRAPHY ; EXTRACTION ; EXPRESSION ; PARTICLES ; MEDIA ; 3A4
Funding ProjectNational Key Research and Development Program of China[2018YFC0311101] ; Beijing Natural Science Foundation[L192017] ; National Natural Science Foundation of China[21821005] ; National Natural Science Foundation of China[22078334] ; National Natural Science Foundation of China[21902160] ; Youth Top Talent Project of Organization Department of Beijing Municipal Committee[ZZB2019001]
WOS Research AreaChemistry
WOS SubjectChemistry, Analytical
Funding OrganizationNational Key Research and Development Program of China ; Beijing Natural Science Foundation ; National Natural Science Foundation of China ; Youth Top Talent Project of Organization Department of Beijing Municipal Committee
WOS IDWOS:000681596600001
PublisherWILEY-V C H VERLAG GMBH
Citation statistics
Document Type期刊论文
Identifierhttp://ir.ipe.ac.cn/handle/122111/49597
Collection中国科学院过程工程研究所
Corresponding AuthorHuang, Yongdong; Ma, Guanghui
Affiliation1.Chinese Acad Sci, Inst Proc Engn, State Key Lab Biochem Engn, Beijing 100190, Peoples R China
2.China Univ Min & Technol, Sch Chem & Environm Engn, Beijing, Peoples R China
3.Beijing Inst Petrochem Technol, Dept Appl Chem, Beijing, Peoples R China
4.Univ Chinese Acad Sci, Sch Chem Engn, Beijing, Peoples R China
Recommended Citation
GB/T 7714
Zhao, Lan,Tao, Jiaoli,Huang, Yongdong,et al. Tailored nanodisc immobilization for one-step purification and reconstitution of cytochrome P450: A tool for membrane proteins' hard cases[J]. JOURNAL OF SEPARATION SCIENCE,2021:12.
APA Zhao, Lan.,Tao, Jiaoli.,Huang, Yongdong.,Zhu, Kai.,Du, Yuxiang.,...&Ma, Guanghui.(2021).Tailored nanodisc immobilization for one-step purification and reconstitution of cytochrome P450: A tool for membrane proteins' hard cases.JOURNAL OF SEPARATION SCIENCE,12.
MLA Zhao, Lan,et al."Tailored nanodisc immobilization for one-step purification and reconstitution of cytochrome P450: A tool for membrane proteins' hard cases".JOURNAL OF SEPARATION SCIENCE (2021):12.
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