角蛋白单体获得及诱导S.maltophilia DHHJ产角蛋白酶研究

A preliminary study on biochemical mechanism of degrading keratins by Stenotrophomonas maltophilia DHHJ

可降解羽毛微生物能以羽毛粉为唯一营养源生长,而羽毛粉是一种大颗粒的不溶性底物,不能直接进入细胞内作为营养源同时作为一级信使来诱导酶基因表达。本文通过化学还原法水解羽毛得到可溶性羽毛角蛋白溶液,利用电泳和质谱验证其分子量约10 k D,为角蛋白单体。分别以该可溶性羽毛角蛋白及角蛋白单体酶解片段为诱导源,在无诱导源、羽毛粉为对照的情况下,测定72 h内嗜麦芽窄食单胞菌(S.maltophilia)DHHJ产生的角蛋白酶的酶活。在无诱导源时,角蛋白酶基因表现本底表达(0.5 U/m L);培养基中添加羽毛粉及角蛋白单体时,角蛋白酶表达量高,分别可达15 U/m L和20 U/m L。并且角蛋白单体酶解片段诱导酶表达较低(2.3 U/m L)。该结果初步表明,水溶性角蛋白单体作为信号源与细菌细胞接触或进入细胞内,控制角蛋白酶基因表达。该结果为细菌降解角蛋白分子机制研究奠定基础。

Feather powder is large and insoluble. It cannot express signals directly to intracellular. The soluble feather keratin was obtained by chemical reduction method to hydrolyze feather,which was monomer with a molecular weight of 10 k D verified by SDS-PAGE and mass spectrometry. The enzyme activities of keratinase produced by Stenotrophomonas maltophilia DHHJ within 72 h were measured under the following 4 conditions： no induced source,soluble feather keratin as induced source,keratin digested fragment as induced source and feather powder as induced source. The enzyme activities of keratinase was quite low without induced source. When the induced source existed,Stenotrophomonas maltophilia DHHJ was induced to produce keratinase. Both the feather powder and soluble feather keratin could induce higher enzyme activities of keratinase（ 15 U / m L and 20 U / m L,respectively）. While the keratin enzymatic fragment could only induce lower enzyme activities of keratinase（ 2. 3 U / m L）.