草菇胞外高丰度蛋白质的分离测序及cDNA片段克隆

Purification of Two High Expressional Exocellular Proteins From Volvariella Volvacea and Cloning one of Cdnd Fragments

经过柱层析,从以稻草粉为基质培养的草菇(Volvariella volvacea)摇瓶发酵液中纯化了HEP1(h ighexpressional protein 1)和HEP2(h igh expressional protein 2)两个在胞外高丰度存在的蛋白质.SDS-PAGE电泳和分子筛层析的结果表明HEP1和HEP2的相对分子质量分别是52.6×103和26.2×103,且都是单亚基.其中HEP1经过酶解印迹后对肽段N端前15个氨基酸进行测序.根据测序结果设计简并引物,PCR扩增到一条长约750 bp特异核酸条带.克隆测序后,该cDNA片段经GenBank多序列比对,未发现其属于某个特定的基因家族.

Two exocellular proteins , HEP1 and HEP2 are abundant in fermentation broth , were purified from straw powder culture of Vovareilla volvacea through protein chromatograph methods . The result of SDS-PAGE and gel filtration indicated that HEP1 and HEP2 are both composed of one polypeptide with the molecular mass of 52.6 × 103 and 26.2 × 103 , respectively . The HEP1s fifteen amino acid residues of N-terminal was determined . Degenerated primers designed according to the N - terminal amino acid , The 750 bp cDNA fragment was amplified with degenerated primers and aligned with GenBank. The orthol- ogous sequences wasnt found.