本文選題：角蛋白酶　切入點(diǎn)：aprA　出處：《現(xiàn)代食品科技》2016年12期

【摘要】：從蛇消化道內(nèi)容物分離出的一株產(chǎn)角蛋白酶菌株為蠟樣芽孢桿菌YSQ08。通過(guò)分析Bacillus cereus ATCC 14579的基因組和蛋白酶家族,對(duì)比分析蠟樣芽孢桿菌YSQ08純化角蛋白酶的酶活特性,確定堿性蛋白酶aprA基因(1194 bp)為目標(biāo)角蛋白酶。為深入研究角蛋白酶的潛在能力,對(duì)蠟樣芽孢桿菌YSQ08堿性蛋白酶aprA基因在畢赤酵母X33上進(jìn)行克隆、表達(dá)和優(yōu)化重組。結(jié)果其10倍濃縮的角蛋白酶酶活達(dá)到122.60 U/mL。經(jīng)優(yōu)化后的aprA基因在表面展示重組酵母中得到高效表達(dá),并且具有較高的角蛋白酶活性,酶活最高可達(dá)到295.78 U/g干細(xì)胞重。酶學(xué)性質(zhì)分析結(jié)果表明,表面展示表達(dá)的重組aprA蛋白酶性質(zhì)與天然型相比具有更高的熱穩(wěn)定性,最適反應(yīng)溫度為55℃,最適pH為8.0。Fe~(2+)可顯著提高重組蛋白的酶活性,最高可達(dá)329.08%。表面展示表達(dá)的重組角蛋白酶可作為全細(xì)胞固定化催化劑,具有更好的熱穩(wěn)定性并能降低酶的純化回收成本。
[Abstract]:A keratinase producing strain isolated from the contents of snake digestive tract was Bacillus cereus YSQ08.By analyzing the genome and protease family of Bacillus cereus ATCC 14579, and comparing the activity of keratinase purified by Bacillus cereus YSQ08, we confirmed that aprA gene of alkaline protease (aprA) was the target keratinase.In order to study the potential ability of keratinase, the YSQ08 alkaline protease aprA gene of Bacillus cereus was cloned, expressed and recombined with Pichia pastoris X33.Results the activity of 10 times concentrated keratinase was 122.60 U / mL.The optimized aprA gene was highly expressed in the surface display recombinant yeast and had high keratinase activity, the highest activity of which could reach 295.78 U / g stem cell weight.The results of enzymatic analysis showed that the properties of recombinant aprA protease expressed on the surface had higher thermal stability than that of the natural type. The optimum reaction temperature was 55 鈩，

本文編號(hào)：1708335