To analyze the bone-inducing activity of C terminal of hBMP-2 and get a new recombinant product of hBMP-2, the gene encoding 102 aa of hBMP-2 mature peptide C terminal was cloned and expressed in E. coli and the first Cys was mutated with Ser. The fragments encoding the target peptide were amplified and cloned into heat-inducible expression vector pDH and transformed into E. coli DH5 alpha. After induction, a new protein bond appeared on the SDS-PAGE. The expressed products amounted to 30% of the total bacterial protein, which existed in the form of inclusion body. The products of bacterial lysates were purified through the ion-exchange chromatography. The denatured proteins were dialysed and diluted directly into the refolding buffer. The renatured products were implanted into mouse thigh muscles to analyze their bone-inducing activity respectively. The results of histological assay showed that the 102 peptide of hBMP-2 could ectopically induce formation of bone, while the mutated 102 peptide of hBMP-2 could not. It suggested hBMP-2 102 peptide still had bone-inducing activity. The first Cys of hBMP-2 mature peptide might be necessary for integrity of three pairs of disulfide bond, and also essential for bone-inducing activity of hBMP-2.
[Bone-inducing activity of human bone morphogenetic protein-2 102 peptide] Publishing Authors By Initials
