title: Complete solubilization and purification of recombinant human growth hormone produced in Escherichia coli.
abstract: High-level expression of recombinant human growth hormone (hGH) in Escherichia coli (E. coli) leads to the formation of insoluble aggregates as inclusion bodies devoid of biological activity. Until recently, significant efforts have been made to improve the recovery of active hGH from inclusion bodies. Here, we developed an efficient procedure for the production of completely soluble hGH by minimizing the formation of inclusion bodies and optimizing protein purification conditions. Under the newly established conditions we were able to obtain most of the total hGH in the soluble fraction. We show that the soluble protein can be efficiently purified in high yield by a series of chromatographic procedures. We analyzed the resulting hGH using various analytical techniques such as reversed-phase high-performance liquid chromatography (RP-HPLC), size-exclusion chromatography (SEC), matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry, and circular dichroism (CD). These multiple analyses support the conclusion that we obtained highly pure hGH with the expected molecular mass and intact secondary structure. The biological activity of purified hGH was also confirmed by evaluating its growth-promoting effect using a cell proliferation assay. Taken together, we describe a straightforward strategy for the production of completely soluble and biologically active hGH in E. coli.
authors: Center for Bioanalysis, Department of Metrology for Quality of Life, Korea Research Institute of Standards and Science, 1 Doryong-dong, Youseong-gu, Daejeon, South Korea.
authors: Min-Ji
authors: MJ
authors: Kim
authors: Hyun Soo
authors: HS
authors: Park
authors: Kyung Hye
authors: KH
authors: Seo
authors: Hyo-Jin
authors: HJ
authors: Yang
authors: Sook-Kyung
authors: SK
authors: Kim
authors: Jun-Hyuk
authors: JH
authors: Choi
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D003593:Cytoplasm / Q000235:genetics; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics*; D005818:Genetic Engineering / Q000379:methods*; D019382:Human Growth Hormone / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification* / Q000494:pharmacology; D006801:Humans; D012995:Solubility; D013696:Temperature
chemical_list: D019382:Human Growth Hormone