摘要
利用RT-PCR从人肥大前列腺组织钓取94个氨基酸的人前列腺分泌蛋白(PSP94)全长cDNA,序列分析结果与文献报道的完全一致.将PSP94成熟肽与人TNFα衍生物(TNFΔ)通过Linker-SAPGTP在基因水平上融合成5′PSP94-TNFΔ,融合基因DNA序列分析结果与设计的相符合.5′PSP94-TNFΔ在大肠杆菌中表达产物分子量约为31kD,表达量约占菌体总蛋白量的35%.以L929细胞和人前列腺癌细胞株PC-3为靶细胞进行细胞毒分析结果表明,5′PSP94-TNFΔ融合蛋白既具有TNF的细胞毒活性。
Human prostate secretory protein of 94 amino acids (PSP94) cDNA including signal and mature peptide has been successfully obtained by reverse transcription polymerase chain reaction (RT PCR) from human hypertrophic prostate mRNA.Sequence analysis indicated that the sequence of human PSP94 cDNA was the same as that of human natural PSP cDNA.The hPSP94 cDNA and human TNFα mutant (TNF Δ) gene were fused by an artificial linker SAPGTP.The chimeric gene coding for 5′PSP SAPGTP TNF Δ fusion protein under control of the phage P RP L promoter was constructed.The expression was assayed by SDS PAGE,which showed that the amount of the expressed the protein was about 35% of total bacterial proteins.Western blot indicated that the molecular weight of 5′PSP SAPGTP TNF Δ was in accordance with the estimated value of 31 kD.Through biological activity analysis,the fusion protein has cytotoxicity activity of L929 and PC 3 cell.
出处
《中国生物化学与分子生物学报》
CAS
CSCD
1999年第3期359-364,共6页
Chinese Journal of Biochemistry and Molecular Biology