Nacre, also known as mother of pearl, contains factors that can promote osteogenesis. In this study, pearlin, a protein from the mantle tissue of the pearl oyster Pinctada fucata, was expressed and purified, and its functional activity analyzed using two mineralogenic cell lines, MRC-5 (fibroblasts) and MC3T3-E1 (preosteoblasts). The open reading frame of pearlin was subcloned into the expression vector pET32a( ) and used to transform Escherichia coli BL21 (DE3) strain. The expression of the recombinant protein (molecular weight: 34.19 kDa) was induced by isopropyl-β-D-thiogalactopyranoside in the form of inclusion bodies which were solubilized in 8 M urea. The recombinant protein was renatured by stepwise dialysis, purified by Ni-NTA affinity chromatography, and analyzed for the effects on cell proliferation by the MTT assay and osteoblastic differentiation by alkaline phosphatase (ALP) activity. The recombinant pearlin promoted proliferation ofMRC-5 andMC3T3-E1 cells at 10 µg/mL and increased their ALP activity at 2.5 µg/mL and 5 µg/mL, respectively. The present study showed that the recombinant pearlin exerted functional effects similar to those of the natural protein, laying a foundation for large-scale production of pearlin in a prokaryotic system.
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Vol. 36 • No. 2