YS-80-122海洋低温碱性蛋白酶的性质

THE CHEMICAL CHARACTERISTICS OF MARINE LOW-TEMPERATURE ALKALINE PROTEASE(YS-80-122)

碱性蛋白酶是一种最重要的工业用酶 ,广泛应用于洗涤、食品、皮革和丝绸行业。海洋低温碱性蛋白酶是一种应用我国海洋嗜极微生物黄海黄杆菌 (YS - 80 - 12 2 )代谢产物并通过现代生物工程手段开发出的一类新型酶。通过 6 0 0 0转 /分离心 (4℃ ) ,收集 (30 % - 6 5 % )硫酸铵沉淀 ,透析后等电聚焦制备分离 (12W) ,并经凝胶S - 2 0 0过滤 ,获得纯品经SDS -PAGE聚丙烯酰胺凝胶和C8HPLC图谱已达到电泳纯。其分子量为 4 9,32 0Dal,等电点为 8.5。建立酶活—温度—pH方程与模型 (r2 >0 .9) ,可求出其最适温度为 30℃ ,最适 pH为 9.5 ,低温 (T≤30℃ )和低 pH(pH≤ 8)对保持其稳定性有利。经试验该酶与大多数金属离子和增稠剂及表面活性剂适应性良好。Pb2 +、Ag+、Cu2 +对其有抑制作用 ,EDTA对其有强烈抑制作用。该酶性质与以往发现碱性蛋白酶性质不同 ,属于一种新型海洋酶。

Alkaline proteases constitute one of the most important groups of industrial enzymes, and play an important role in the detergent, food, pharmaceutical, leather and silk industries. Marine Low temperature Alkaline Protease(MLAP) is a kind of extracellular enzyme from a marine bacterium Flavobacterium YS-80-122. Purification is carried out by centrifuging the fermentation medium at a high speed (6000r/m) and low temperature(4℃) and by precipitating by ammonium sulphate (30%-65%). The dialyzate was applied to an IEF producing system(12W) and column Sephacry I S-200 . Finaly we obtained the electrophoreticly pure enzyme and the purity was checked by SDS PAGE and C8 HPLC. Its molecular weight is 49,320Dal and its pH 8.5. The model of enzyme activity temperature pH ( r 2>0.9) was made by studying the effects of temperature and pH on enzyme activity and stability. Its optimum pH is 9.5, temperature optima at 30℃. The enzyme is stable at low temperature(T≤30℃)and lower pH (pH≤8)is helpful for keeping its stability. And protease is compatible with most of the metal ions and surfactants. few Metal ions Pb 2+ , Ag + and Cu 2+ can act as a strong inhibitor of its activity. Strong inhibition was observed with EDTA, and the finding was different from former alkaline proteases.