Expression, purification, and characterization of a novel soluble form of human Delta-like-1.
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 6x His tag, was expressed in Escherichia coli as inclusion bodies after isopropyl beta-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 microg/ml of rhDSL. The rhDSL protein might therefore be a potential supplement for the expansion of UCB-primitive hematopoietic cells.
Zhao
Mei
M
Laboratory of Regenerative Medicine, School of Pharmacy, Shanghai Jiao Tong University, Shanghai 200240, People's Republic of China.
Wu
Mingyuan
M
Guo
Lingchen
L
Jiang
Junfen
J
Huang
Weiwei
W
Lin
Xiaojuan
X
Zhang
Zhonghui
Z
Xiang
Di
D
Lu
Huili
H
Zhu
Shunying
S
Yu
Yan
Y
Moldenhauer
Anja
A
Han
Wei
W