在黄孢原毛平革菌（Phanerochaete chrysosporium）的限氮浸没式培养中，研究了不同条件对选择性合成木质素过氧化物酶（LiP）和锰过氧化物酶（MnP）的作用。LiP只在很窄的氮源浓度范围内（0.8～1.8mmol/L）才能测到，而MnP在较宽浓度范围内（0.4～1.8mmol/L）表现出较高的酶活水平.培养基中缺乏Mn2+不能合成MnP，但可以得到活力较高的LiP。[Mn2+]在0.06～0.84mmol/L时可获得LiP。添加微量Mn2+即可获得较高活力的MnP，此后增加Mn2+对MnP的活力影响不大，直至浓度为3.36mmol/L才表现出明显的抑制作用。通纯氧可使LiP活力提高50%，但对MnP影响不大。

在黄孢原毛平革菌（Phanerochaete chrysosporium）限氮浸没式培养中，获得了含木质素过氧化物酶（LiP）和锰过氧化物酶（MnP）、只含LiP或MnP以及无LiP和MnP 的4种培养物，考察了LiP，MnP及菌球在染料脱色过程中的作用。结果表明，4 种培养物对6种染料都有明显的脱色效果，甲基橙、橙I、次甲基蓝脱色率在90%左右，刚果红、直接湖蓝脱色率在80%左右，结晶紫脱色率在60%左右。无酶情况下染料的脱色主要是菌球的吸附，有酶情况下染料的脱色主要是酶的降解。菌球在脱色过程中除了吸附染料外，还起到提供H2O2和藜芦醇的作用，促进了染料的降解。

比较了黄孢原毛平革菌在3 种生物反应器（搅拌式反应器、鼓泡式反应器、曝气式反应器）中合成木质素过氧化物酶（LiP）和锰过氧化物酶（MnP）的差异。结果表明，曝气式反应器对酶的合成（尤其是LiP）最为有利. 考察了曝气式反应器中半连续培养和连续培养两种方式下酶的合成和橙I脱色情况，发现半连续培养可使培养体系长时间保持较高酶活力，置换比例为1/2时染料废水可连续脱色5批，脱色率达到90%以上，比脱色率在46.7g/（g×d）以上。连续培养条件下酶很快失活，废水的脱色率迅速下降。在曝气式反应器中用半连续培养的方式（置换比例1/2）对实际印染废水进行处理，可处理废水4批，前3批脱色率达到90%以上，第4批有明显下降。

Abstract A b-N-acetylglucosaminidase gene (nag3A) from Clostridium paraputrificum M-21 was cloned in Escherichia coli. The nag3A gene consists of an open reading frame of 1,239-bp, encoding 413 amino acids with a deduced molecular weight of 45,531 Da. Nag3A is a single domain enzyme containing a family 3 glycoside hydrolase catalytic domain. Nag3A was purified from recombinant E. coli and characterized. The enzyme hydrolyzed chitooligomers such as di-N-acetylchitobiose, tri-N-acetylchitotriose, tetra-N-acetylchitotetraose, penta-N-acetylchitopentaose, hexa-N-acetylchitohexaose, ballmilled chitin, and synthetic substrates such as 4-methylumbelliferyl N-acetyl b-d-glucosaminide [4-MU-(Glc-NAc)], but had no activity at all against p-nitrophenyl-bd-glucoside, p-nitrophenyl-b-d-xyloside, or p-nitrophenyl-b-d-galactosamine. The enzyme was optimally active at 50℃ and pH 7.0, and the apparent Km and Vmax values for 4-MU-(GlcNAc) were 7.9μM and 21.8μmol min-1mg protein-1, respectively. SDS-PAGE, zymogram, and immunological analyses suggested that this enzyme is induced by ball-milled chitin.

A fi-N-acetylglucosaminidase gene (nag84A) was cloned from Clostridium paraputr$cum M-21 in Escherichia coli. The nag84A gene consists of an open reading frame of 4647 bp encoding 1549 amino acids, with a deduced molecular weight of 174,311, which have a catalytic domain belonging to family 84 of the glycoside hydrolases. Nag84A was purified from a recombinant E. coli and characterized. Although Nag84A exhibited high homology to the hyaluronidase from Clostridium perfringens, it did not degrade hyluronic acid. The enzyme hydrolyzed chitooligomers such as di-, tri-, tetra-, penta- and hexa-N-acetylchitohexaose, and synthetic substrates such as 4-methylumbelliferyl N-acetyl fi-D-glucosaminide (4-MU-(GlcNAc)], but did not hydrolyze 4-MU-fl-D-glucoside,4-MU-a-D-glucoside, 4-MU-a-D-GlcNAc, 4-MU-a-D-galactoside, 4-MU-fl-D-xyloside, PNPP-D-galactoside, and PNP-a-D-xyloside. The enzyme was optimally active at 5O℃ and pH 6.5, and the apparent K, and Vm,, values for 4-MU-(GlcNAc) were 8.5pM and 1.39 ymol/min/mg of protein, respectively. SDS-PAGE, zymogram, and immunological analyses suggested that Nag84A was inducible by ball-milled chitin. Since Nag84A has a high molecular weight with a family 84 catalytic domain with high homology to hyaluronidases but no hyaluronidase activity, the enzyme is a novel j3-N-acetylglucosaminidase different from others reported having low molecular weights and belonging to family 3 and family 18.