Human CD49f (Integrin a6) has been demonstrated to be specific membrane protein marker for human hematopoietic stem cell (HSC). Two peptide AG-10 (NRWHSIYITRFG) and AG-32 (TWYKIAFQRNRK), which derived from human laminin-111, has been reported to engage with the α6β1 integrin very specifically. To generate a synthetic matrix protein for coating to maintain HSC culture in vitro, AG-10 and AG-32 peptide was fused to human Vitronectin (VNT) C-terminal as recombinant protein. This recombinant VNT-AG10+32 fusion protein may be served as synthetic matrix protein for interaction with α6β1 integrin positive cells.

DNA fragment which encoded for two peptide AG-10 (NRWHSIYITRFG) and AG-32 (TWYKIAFQRNRK) was fully synthesized and fusion to human vitronectin (63-398aa) C-terminal with flexible linker domain (GGGGSGGGGS) for generating VNT-AG10+32 fusion protein. VNT-AG10+32 DNA fragment was constructed with codon optimization and expressed in E.coli as inclusion bodies. The final product was refolded using our unique “temperature shift inclusion body refolding” technology and chromatographically purified.