title: High-level expression and purification of the major house dust mite allergen Der p 2 in Escherichia coli.
abstract: Der p 2, a major allergen derived from the house dust mite Dermatophagoides pteronyssinus, is one of the most clinically relevant allergens worldwide. Recombinant Der p 2 (rDer p 2) is useful in clinical diagnosis and disease-specific immunotherapy. However, previous studies showed that Der p 2 can only be expressed in Escherichia coli (E. coli) cells as inclusion bodies, thus protein refolding is required to obtain functional products. Here we report a new method to produce biologically active Der p 2 protein in E. coli. N-terminal hexahistidine- and trigger factor (TF)-tagged Der p 2 was expressed in soluble form in E. coli and purified using a combination of chromatography processes. This procedure produced milligram-level high purity Der p 2 per liter of bacterial culture. Moreover, far-UV region circular dichroism (CD) analysis and serum specific IgE reactivity test demonstrated that the secondary structure and IgE reactivity properties of rDer p 2 produced in our study were almost identical to those of natural Der p 2 (nDer p 2). In conclusion, the method developed in this work provides a useful tool for the production of immunologically active recombinant Der p 2 for clinical applications.
authors.affiliation: Department of Clinical Laboratory, Peking University Shenzhen Hospital, Shenzhen, 518036, Guangdong, China; Department of Dermatology, Peking University Shenzhen Hospital, Shenzhen, 518036, Guangdong, China; Shantou University Medical College, Shantou, 515041, Guangdong, China.
authors.forename: Tuo
authors.initials: T
authors.lastname: Cao
authors.affiliation: Department of Dermatology, Peking University Shenzhen Hospital, Shenzhen, 518036, Guangdong, China; Shenzhen Key Lab for Translational Medicine of Dermatology, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China.
authors.forename: Zhang
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Biomedical Research Institute, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China.
authors.forename: Zhi-Gao
authors.initials: ZG
authors.lastname: Liu
authors.affiliation: Department of Dermatology, Peking University Shenzhen Hospital, Shenzhen, 518036, Guangdong, China; Shenzhen Key Lab for Translational Medicine of Dermatology, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China.
authors.forename: Xia
authors.initials: X
authors.lastname: Dou
authors.affiliation: Department of Dermatology, Peking University Shenzhen Hospital, Shenzhen, 518036, Guangdong, China; Shenzhen Key Lab for Translational Medicine of Dermatology, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Biomedical Research Institute, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China; Shenzhen Key Lab for Translational Medicine of Dermatology, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Department of Dermatology, Peking University Shenzhen Hospital, Shenzhen, 518036, Guangdong, China; Shenzhen Key Lab for Translational Medicine of Dermatology, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Wu
authors.affiliation: Central Laboratory, Peking University Shenzhen Hospital, Shenzhen, 518036, Guangdong, China.
authors.forename: Zhen-Dong
authors.initials: ZD
authors.lastname: Yu
authors.affiliation: Biomedical Research Institute, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China; Shenzhen Key Lab for Translational Medicine of Dermatology, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China. Electronic address: biowei@163.com.
authors.forename: Zhun
authors.initials: Z
authors.lastname: Wei
authors.affiliation: Department of Dermatology, Peking University Shenzhen Hospital, Shenzhen, 518036, Guangdong, China; Shenzhen Key Lab for Translational Medicine of Dermatology, Shenzhen Peking University - the Hong Kong University of Science and Technology Medical Center, Shenzhen, 518036, Guangdong, China. Electronic address: yubomd@hotmail.com.
authors.forename: Bo
authors.initials: B
authors.lastname: Yu
mesh_terms: D000818:Animals; D039741:Antigens, Dermatophagoides / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D060829:Arthropod Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression / Q000276:immunology; D006801:Humans; D017433:Protein Structure, Secondary; D039981:Pyroglyphidae / Q000276:immunology* / Q000472:pathogenicity; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification
chemical_list: D039741:Antigens, Dermatophagoides; D060829:Arthropod Proteins; C451163:Dermatophagoides pteronyssinus antigen p 2; D011994:Recombinant Proteins
keywords: Allergy; Der p 2; House dust mite; Recombinant protein