CELLOBIOSE DEHYDROGENASES OF SPOROTRICHUM-(CHRYSOSPORIUM)-THERMOPHILE
CELLOBIOSE DEHYDROGENASES OF SPOROTRICHUM-(CHRYSOSPORIUM)-THERMOPHILE
复制标题
DOI：
10.1111/j.1432-1033.1991.tb15984.x
复制
发表时间：
1991-05-23
期刊：
影响因子：
--
通讯作者：
DREYER, JL
中科院分区：
文献类型：
--
作者：
CANEVASCINI, G;BORER, P;DREYER, JL
经DEAE-Trisacryl柱层析分离得到的嗜热芽孢霉纤维二糖脱氢酶ATCC 42464，经不同的层析技术纯化后，分别命名为纤维二糖脱氢酶I和纤维二糖脱氢酶II。这两种酶都是轻微糖基化的黄色素-b蛋白，具有相似的催化性质，但具有不同的分子质量(酶I和酶II分别为91 kDa和192 kDa)和等电点(4.1对3.45)。SDS/PAGE分析表明，较大的酶II为同源二聚体，其亚基与脱氢酶I相近，但不同于脱氢酶I。在用木瓜酶有限地消化这两种酶之后，形成了两个具有残留活性的主要组分，一个含有血红素，另一个是黄素成分；每个组分都有其独特的层析行为。黄素携带组分显示非典型的(对于黄素蛋白而言)三条带光谱，表明黄素衍生物的存在。这两种酶与氧的反应都非常缓慢，显然会形成一些超氧自由基，可能还会产生过氧化氢。纤维二糖和其他纤维糊精在其还原糖基部分被氧化成相应的醛酸。使用可自氧化的吩嗪亚硫酸盐，纤维素(水合形式或结晶形式)也会在自由还原末端被氧化，从而在酶解时释放出相当数量的纤维二酸。
Both cellobiose dehydrogenases of Sporotrichum (Chrysosporium) thermophile, ATCC 42464, obtained after fractionation with DEAE-Trisacryl chromatography and named cellobiose dehydrogenase I and II have been purified to homogeneity by different chromatographic techniques. Both enzymes are slightly glycosylated flavocytochrome-b proteins with similar catalytic properties but with distinct molecular masses (91 kDa and 192 kDa for enzymes I and II, respectively) and isoelectric point (4.1 versus 3.45). Examination by SDS/PAGE clearly showed that the larger enzyme II is a homodimer, whose subunit is close to, but different from dehydrogenase I which is homogeneous by this technique. After limited digestion of both enzymes with papain, two main fractions with residual activity are formed, one carrying the heme, the other being the flavin component; each fraction is characterized by its particular chromatographic behaviour. The flavin carrying component shows an atypical (for flavoprotein) three-banded spectrum indicative of the presence of a flavin derivative. Both enzymes react very slowly with oxygen clearly forming some superoxide radicals and possibly hydrogen peroxide. Cellobiose and other cellodextrins are oxidized at their reducing glycosyl moiety to the corresponding aldonic acid. With the use of the autooxidable phenazinemethosulphate, cellulose (either in a hydrated form or crystalline) is also oxidized at free reducing ends so that appreciable amounts of cellobionic acid are released upon enzymatic hydrolysis.