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Expression, Purification, and Characterization of a Novel Soluble Form of Human Delta-like-1

Zhao, Mei, Wu, Mingyuan, Guo, Lingchen, Jiang, Junfen, Huang, Weiwei, Lin, Xiaojuan, Zhang, Zhonghui, Xiang, Di, Lu, Huili, Zhu, Shunying, Yu, Yan, Moldenhauer, Anja, Han, Wei
Applied biochemistry and biotechnology 2010 v.160 no.5 pp. 1415-1427
Escherichia coli, affinity chromatography, anion exchange, cell differentiation, humans, inclusion bodies, signal transduction, stem cells, umbilical cord
The notch signaling pathway plays an important role in inhibiting cell differentiation and enhancing the repopulation capability of hematopoietic stem/progenitor cells. In this study, we developed rhDSL, a novel soluble form of Notch ligand Delta-like-1, which contains the DSL domain and the N-terminal sequence of the ligand, and investigated its function in ex vivo expansion of human umbilical cord blood (UCB)-primitive hematopoietic cells. The coding sequence for rhDSL was cloned into a pQE30 vector, and the recombinant rhDSL, fused with a 6× His tag, was expressed in Escherichia coli as inclusion bodies after isopropyl β-d-thiogalactoside induction. After renaturing by dilutions, the protein was purified through anion exchange followed by affinity chromatography. The purity of rhDSL protein was more than 99% with very low endotoxin. In combination with human c-kit ligand, the effect of rhDSL on ex vivo expansion of UCB CD34⁺ cells was found to be optimal at 1.5 μg/ml of rhDSL. The rhDSL protein might therefore be a potential supplement for the expansion of UCB-primitive hematopoietic cells.