本文選題：雅致放射毛霉 + 羧肽酶Y��； 參考：《現(xiàn)代食品科技》2017年01期

【摘要】：本研究從雅致放射毛霉中克隆、表達(dá)和純化羧肽酶Y(CPY),體外激活后檢測(cè)其酶學(xué)性質(zhì)。本文篩選并鑒定了一株腐乳發(fā)酵菌株,命名為雅致放射毛霉PEP001(Actinomucor elegans PEP001)。以相近種族來(lái)源的真菌羧肽酶Y保守序列設(shè)計(jì)簡(jiǎn)并引物調(diào)取部分目的基因序列,利用末端擴(kuò)增技術(shù)(RACE)分別獲得3’與5’端基因全長(zhǎng),克隆獲得羧肽酶Y基因,全長(zhǎng)為1557 bp,編碼518 aa。將其酶原(pro CPY)在大腸桿菌Rosetta中重組表達(dá),產(chǎn)物為包涵體;在畢赤酵母GS115中成功分泌表達(dá)pro CPY,產(chǎn)量為151.20±10.20mg/L。通過(guò)體外激活與純化,獲得電泳純的成熟羧肽酶Y。成熟羧肽酶Y最適pH為6.0,最適溫度為45℃,在40℃下有較高穩(wěn)定性,在60℃下很快失活,在pH 4.0~7.0下都有較高的穩(wěn)定性。本文中雅致放射毛霉PEP001羧肽酶Y為首次報(bào)道,為進(jìn)一步研究其應(yīng)用打下一定基礎(chǔ)。
[Abstract]:In this study, we cloned, expressed and purified carboxypeptidase Yapeptidase from Mucor radiorrhoeae, and detected its enzymatic properties after activation in vitro. In this paper, a sufu fermentation strain was screened and identified, named PEP001, Actinomucor elegans PEP001. A degenerate primer was designed with conserved sequence of fungal carboxypeptidase Y from similar ethnic origin to obtain partial target gene sequence. The complete length of 3 'and 5' terminal genes were obtained by terminal amplification technique, and the carboxypeptidase Y gene was cloned. The total length is 1557 BP, encoding 518 aa. The enzyme was expressed in Escherichia coli Rosetta and secreted successfully in Pichia pastoris GS115 with a yield of 151.20 鹵10.20 mg / L. The mature carboxypeptidase Y. was obtained by in vitro activation and purification. The optimum pH of mature carboxypeptidase Y is 6.0, the optimum temperature is 45 鈩，

本文編號(hào)：2016486