一种中性普鲁兰酶及其在红薯粉丝制作中的应用

Characterization of a Neutrophil Pullulanaseand Its Application in Sweet Potato Vermicelli Processing

摘要:

本研究旨在开发一种在较宽泛的pH范围内活性稳定的中性普鲁兰酶，以适应以红薯面为原料的粉丝制作工艺。以地表地芽孢杆菌（Geobacillus subterraneus） XQ3665基因组DNA为模板，经PCR扩增得到普鲁兰酶基因pul3665，并实现了异源表达。重组酶PUL3665纯酶比酶活为32 U/mg，最适反应温度55 ℃，最适pH为6.5，在pH 5.5~7.5 范围内酶活保持在最高酶活的80%以上。PUL3665降解普鲁兰多糖的产物为麦芽三糖，不降解可溶性直链淀粉，是一种I型普鲁兰酶。在以不同pH值的红薯面为原料的粉丝制作工艺中，按每克芡糊淀粉2 U的量添加枯草芽孢杆菌表达的PUL3665粗酶液处理芡糊，均能在不添加明矾或其它添加剂的条件下制作出粉丝，且成品粉丝质量与添加明矾获得的粉丝基本一致。PUL3665有希望开发成一种用量小，使用效果稳定的新型明矾替代物。

Abstract:

This research aimed to develop a neutral pullulanase with stable activity in a wide pH range, which can adapt to the manufacturing process of vermicelli with sweet potato powder as raw material. The pullulanse-encoding gene pul3665 was cloned from genome DNA of Geobacillus subterraneus XQ3665 by PCR and heterologously expressed. Specific activity of the purified recombinant enzyme PUL3665 is 32 U/mg. The optimal temperature and pH of PUL3665 were 55 ℃ and 6.5, respectively. The PUL3665 maintain 80% of its highest activity in the range of pH 5.5~7.7. The product of enzymatic degradation of pullulan was maltotriose, and the PUL3665 did not degrade amylose. Thus, the PUL3665 belongs to type I pullulanases. The crude enzyme of PUL3665 expressed by B. subtilis was used in the traditional vermicelli-making process. The paste was treated with PUL3665 at a dose of 2 U/g of paste. Vermicellis were successfully achieved using different kinds of sweet potato powder as raw materials. Neitheralum nor other additives were used. The quality of the finished vermicelli made from PUL3665 treated paste was about the same to that of the vermicelli which was made with alum as aditives. The PUL3665 is expectedto be developed into a highly effective and reliable substitute of alum in vermicelli production.