Studies on biodegration mechanism of cellulose by white-rotting fungi.

生长在纤维素上的白色木腐菌黄孢原毛平革菌产生氧化还原酶纤维二糖脱氢酶(CDH)。本研究探讨了CDH在纤维素降解中的作用。CDH只在纤维素的无定形表面发生吸附，而在结晶表面不发生吸附，这表明CDH的作用不是直接导致纤维素晶体结构的分解。在纤维素上生长的黄孢霉静态培养中，CDH的大部分活性存在于菌丝体-纤维素复合体中，而不是在培养液中释放。此外，CDH在纤维素酶表面的定位是纤维素酶作用形成的裂隙所特有的。为了获得大量的CDH，对培养条件进行了优化。在培养基中加入小牛血清可以显著提高胞外液中CDH的产量。研究了微晶细菌纤维素分解过程中纤维二糖水解酶I(CBH I)与CDH之间的相互作用，结果表明CDH氧化还原系统通过解除产物抑制而增强了CBH I的活性，并探讨了CDH中黄素与血红素辅因子之间电子传递的pH依赖性机制。结果表明，这种现象是由黄素结构域的蛋白质结构随pH变化而引起的。

The white wood-rotting fungus Phanerochaete chrysosporium grown on cellulose produces the oxidoreductase cellobiose dehydrogenase (CDH). In this study, the role of CDH in cellulose degradation was investigated.Adsorption of CDH is occurred only on the amorphous surface of cellulose, but not on the crystalline surface, suggesting that the function of CDH is not directly responsible for decomposition of the crystalline structure of cellulose.In the static cultures of P.chrysosporium grown on cellulose, most of CDH activity is observed in the mycelium-cellulose complex, but not released in the culture solution. Moreover, localization of CDH on the cellulose surface is specialized to the crack formed by the action of cellulases. This observation is a good evicdence to suggest the cooperative function between cellulases and CDH in vivo.The optimization of cultivation condition has been elaborated to obtain a large amount of CDH.The addition of calf bovine serum in the culture medium is resulted in the significant enhancement of CDH production in the extracellular solution. To investigated the reason of this phenomenon, the method using RT-PCR for measurement of the corresponding mRNA level have been established.The interaction between cellobiohydrolase I (CBH I) and CDH in the decomposition of the microcrystalline bacterial cellulose was investigated The result demonstrated that the CDH redox system enhances CBH I activity by relieving the product inhibition.The mechanism on pH dependence of electron transportation between flavin and heme cofactors in CDH was investigated. The result suggests that this phenomenon is caused by the pH depending conformational changes of the protein structure of the flavin containing domain.