[Molecular cloning and expression of anti-tumor adhesion peptide (beta3)].
To block tumor cell adhesion, inhibit tumor metastasis and recurrence, the anti-adhesion peptide-trimeric beta peptide (DLYYLMDLSYSMKGGDLYYLMDLSYSMKGGDLYYLMDLSYSMK, beta3) was designed. The DNA fragment of beta3 was cloned into expression vector pET-His and the fusion protein His-beta3 was expressed in E. coli. BL21(DE3)plysS. After 1.5 hours' induction with IPTG, His-beta3 peptide was expressed significantly amounting to 10% of the insoluble proteins and 4% of the total proteins. 20mg of beta3 peptide was obtained from one litter culture medium after purification by using metal-chelating sepharose 6B FF. The purity of beta3 is 92.2% according to Gel-Pro analysis. The anti-adhesion effects of beta3 peptide, beta1 peptide (DLYYLMDLSYSMK) and GRGDS on the hepatocellular carcinoma cell line SMMC-7721 and the high metastasis hepatocellular carcinoma cell line HCCLM6 were studied. The result showed the beta3 blocked the adhesion of HCCLM6 cells and SMMC-7721 cells to fibronectin (FN) specifically. The inhibition effect was dose-dependent and time-dependent and the inhibition rate of beta3 was higher than three times concentration of beta1 and GRGDS. This suggested that pET-His-beta3/BL21(DE3)plysS was a suitable expression system for beta3, and the expressed beta3 specially inhibited the adhesion of cancer cells.
Wang
Song-Mei
SM
Laboratory of Molecular Biology, Shanghai Medical College, FuDan University, Shanghai 200032, China.
Zhu
Jun
J
Li
Yan
Y
Pan
Luan-Feng
LF
Zha
Xi-Liang
XL
Liu
Yin-Kun
YK
D000595:Amino Acid Sequence; D000970:Antineoplastic Agents / Q000494:pharmacology*; D002448:Cell Adhesion / Q000187:drug effects; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000494:pharmacology; D010455:Peptides / Q000235:genetics* / Q000378:metabolism* / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D010446:Peptide Fragments; D010455:Peptides; D011994:Recombinant Proteins