漆酶高产菌株的诱变选育及其产酶条件

Mutational screening of fungal strain with high yield of laccase and enzyme production condition of the mutant

以粗毛栓菌Trametesgallica为出发菌,通过紫外诱变处理其担孢子、PDA-RBBR平板变色法初筛、ABTS法测定培养液漆酶酶活力复筛,获得1株漆酶高产诱变菌株SAH-12。用高氮低碳无机盐培养液(LM3)培养时,其峰值酶活力比出发菌株高出4倍,达到5002.6U/L,且产酶稳定。对SAH-12液体培养产酶条件的研究表明:以纤维二糖和蔗糖为碳源明显优于麦麸、淀粉和葡萄糖,其最高酶活分别达18526U/L和13436U/L;有机氮源较无机氮源更有利于SAH-12漆酶的分泌,以蛋白胨、大豆粕和胰化蛋白胨为氮源时其峰值酶活分别达到20544U/L、19671U/L和16180U/L;适宜初始培养pH为4.0;ABTS、单宁酸、没食子酸对产酶均有明显的诱导作用,其中ABTS和单宁酸的诱导效果相对更好,愈创木酚和吐温80对产酶有一定的抑制作用。

The basidiospores of Trametes gallica were exposed under ultra-violet ray and the mutants were screened in RBBR-containing PDA plates, and then the laccase activities of the mutated strains were determined by ABTS. A mutant SAH-12 with high yield of laccase was obtained. The maximum activity of laccase fi＇om SAH-12 had increased up to 5002.6 U/L, and its ability of producing laccase was stable on culture condition of inorganic salt with high nitrogen and low carbon （LM3）. Study of laccase production in the liquid culture of SAH-12 showed that cellobiose and sucrose as carbon source were obviously superior to wheat bran, starch and glucose, and the highest laccase activities were 18526 U/L and 13436 U/L, respectively. Organic compound as carbon source was more suitable for the laccase secretion fi＇om SAH-12 than inorganic compound. Using peptone, soybean meal and tryptone as nitrogen source, the maximum laccase activities were 20544 U/L, 19647 U/L and 16180 U/L, respectively. The initial pH for producing laccas~ fi＇om SAH-12 was 4.0. The ability of producing laccase fi＇om SAH-12 was clearly increased by ABTS, tannic acid and gallic acid, among which ABTS and tannic acid were more superior to gallic acid. Laccase production was inhibited by guaiacol and tween 80.