title: [Expression and purification of recombinant human interleukin 4 in Escherichia coli].
abstract: Human interleukin 4 (IL-4) cDNA was optimized and synthesized according to E. coli preferred codon. A recombinant expression plasmid pET-30a (+)/rhIL-4 was constructed with the target cDNA inserted between Nde I and EcoR I sites, which can translate the mature IL-4 protein with an extra methionine residue at N-terminal. The expression vector was transformed into E. coli BL21 (DE3). The rhIL-4 protein was expressed in the inclusion body. By using the optimized fermentation conditions, the high expression level was achieved with the expression level as high as 35% of total protein obtained. A purification strategy has been designed which includes Q-Sepharose and SP-Sepharose ion-exchange chromatography and dialysis renaturation. The rhIL-4 was purified with the purity more than 98% and the yield of 40 mg per liter fermentation culture achieved. Western blot proved that the purified protein is IL-4. Amino acid sequencing revealed that N-terminal 16 residue sequence is identical to the theoretical sequence. Biological activity assay on TF-1 cells demonstrated that the rhIL-4 is active with an activity of 2.5 x 10(6) AU/mg. This study promises large scale production of rhIL-4.
authors.affiliation: College of Life Science, Fujian Normal University, Fuzhou 350007, China. qybio@163.com
authors.forename: Yan
authors.initials: Y
authors.lastname: Qiu
authors.forename: Jiu-Ru
authors.initials: JR
authors.lastname: Sun
authors.forename: Yang-Bin
authors.initials: YB
authors.lastname: Huang
authors.forename: Zhi-Hua
authors.initials: ZH
authors.lastname: Huang
authors.forename: Lin-Jie
authors.initials: LJ
authors.lastname: Chen
authors.forename: Yong
authors.initials: Y
authors.lastname: Liu
authors.forename: Yue-Xin
authors.initials: YX
authors.lastname: Lin
mesh_terms: D000595:Amino Acid Sequence; D015153:Blotting, Western; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005285:Fermentation; D015870:Gene Expression; D006801:Humans; D015847:Interleukin-4 / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D015847:Interleukin-4