Soluble expression, purification and bioactivity of hemangiopoietin protein
10.3321/j.issn:1007-8738.2009.11.008
- VernacularTitle:HAPO蛋白的可溶性表达、纯化及生物学活性测定
- Author:
Bin LI
;
Xiaojing WANG
;
Yongjun LIU
;
Zhongchao HAN
;
Tianxiang PANG
- Publication Type:Journal Article
- Keywords:
hemangiopoietin(HAPO);
soluble expression;
isolation and purification;
human umbilical vein endothelial cells (HUVEC)
- From:
Journal of Cellular and Molecular Immunology
2009;25(11):991-993,997
- CountryChina
- Language:Chinese
-
Abstract:
AIM: To prepare a soluble hemangiopoietin(HAPO) protein and to construct pET22b(+) expression vector, to obtain pure recombinant HAPO protein and to measure its bioactivity. METHODS: HAPO cDNA was amplified using RT-PCR method from a commercial human fetal liver cDNA library. The resulting product was cloned into pET22b(+) vector and transformed into E.coli BL21(DE3). The recombinant protein was isolated and purified by Ni~(2+)-NTA chelating resin and the chromatographies of SP Sepharose FF. The adhesion of human umbilical vein endothelial cells (HUVEC) were measured by adhesion assay. RESULTS: HAPO gene with a reading frame of 897 bp was successfully cloned from human fetal liver cDNA library, the expressed pET22b(+)-HAPO fused protein existed in a soluble form, with the yield above 10% total bacterial protein and its purity achieved above 80%. The activity assay showed that the treatment of HAPO enhanced total adherence of HUVEC in a concentration-dependent manner. CONCLUSION: HAPO protein can be expressed in a soluble form. HAPO may facilitate the homing of hematopoietic stem/progenitor cells in vitro.
